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Chem PPT Flashcards Unit 2

Mass spectrometry is: The study of matter through the formation of gas phase ions that are characterized using mass spectrometers by their mass, charge, structure, and/or physico-chemical properties

A mass spectrometer is: An analytical instrument that first ionizes a target molecule and then separates and measures the mass-to-charge (m/z) ratio of these molecules or their fragments

A mass spectrometer consists of the following five elements:

Ion souce, vacuum system, mass analyzer, detector, computer

A vacuum system is used to prevent collision of ions during interaction with:

Magnetic or electrical fields

It requires the use of a vacuum from 10-3 to 10-9

torr one of two sources:Mechanical vacuum or efficient high vacuum pump

Class of spectrometer where the ions make one trip through the instrument and then strike the detector, where they are destructively detected is

Beam type

3 types of the above: Quadrupole mass spectrometer, magnetic sensor mass spectrometer, time of flight mass spectrometer

Trapping mass spectrometers: Are based on the trapping of ions to capture and hold ions for an extended time in a small region of space

What does MS/MS stand for? Mass spectrometry/mass spectrometryWhat is also called MS/MS? Tandem mass spectrometryWhat is characterized by a highly selective detection technique used for quantitative analyses, characterization and compound identification?

Tandem mass spectrometry

What does GC-MS stand for? Gas chromatography mass spectrometryWhat is used by the NIST as a definitive method of qualifying standard reference materials and assigning certified values to many clinical analytes?

GC-MS

What is used as confirmatory tests for drug testing presumptively found to be positive by immunochemical analyses?

GC-MS

What does LC-MS stand for? Liquid chromatography mass spectrometryWhat is used for analysis of polar and non polar analytes and large molecular weight compounds?

LC-MS

What is used in screening and confirmation of genetic disorders and inborn errors of metabolism?

LC-MS

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What does MALDI-TOF-MS stand for? Matrix-assisted laser desorption/ionization – Time-of-flight mass spectrometry

What is MALDI-TOF-MS used for? For the detection of specific compounds, identification of proteins, or identification of an organism

What are proteins with catalytic properties? EnzymesWhat are protein catalysts of biological origin? EnzymesWhat is a substance that modifies and increases the rate of a chemical reaction without being permanently changed or consumed?

Catalyst

How are individual enzymes identified? By using the name of the substrate or group upon which they act and then adding the suffix -ase.

The type of reaction involved is identified when naming enzymes.

TRUE

What are the different structures of proteins? Primary, secondary, tertiary, quaternaryBiological activity is affected by denaturization due to what factors?

Elevated temperatures, extremes of pH, changes in ionic strength, and chemical addition

What is one of a group of enzymes catalyzing the same reaction by different molecular structures and characterized by varying physical, biochemical, and immunologic properties?

Isoenzyme

What is a substance that modifies and increases the rate of a particular chemical reaction without being consumed or permanently altered?

A catalyst

What are protein catalyst of biological origin? EnzymesWhat can enzymes convert? Enzymes can convert substrate molecules that

can be used for the measurement of their activity

What defined as the quantity of enzyme that catalyzes the reaction of 1 µmol of substrate per minute?

International unit (IU)

What is the International System of Units (SI)- derived unit for catalytic activity?

Katal unit (KU)

What unit defined moles converted per second? Katal unit (KU)How enzyme concentration governs the rate of Enzyme-Catalyzed Reactions?

The rate of reaction is generally proportional to the amount of enzyme present in the system and is the basis for the quantitative determination of enzymes by measurement of reaction rates.

What is substrate concentration? A reactant in an enzyme-catalyzed reaction that binds to the active center of an enzyme.

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What is first order kinetics? in an enzymatic reaction, as more substrate is consumed, the reaction rate declines and becomes dependent on substrate concentration; this is a phrase of first-order dependence on substrate concentration when the rate of the reaction is proportional to the concentration of substrate or when the enzyme concentration is fixed and the substrate concentration is varied.

What is zero-order kinetics? In an enzymatic reaction, when the reaction rate is constant at maximum value, the reaction rate depends only on enzyme concentration and is independent of substrate concentration; the rate of reaction is proportional to the zero power of the substrate concentration.

What is a constant for a given enzyme acting under given conditions; the experimentally determined substrate concentration at which the enzymatic reaction velocity equals ½ of the maximum velocity of the enzymatic reaction?

Michaelis-Menten constant (Km)

What is a plot of the reciprocal of the velocity of an enzyme catalyzed reaction (ordinate; y-axis) against the reciprocal of the substrate concentration (abscissa; x-axis)?

Lineweaver-Burk plot

What shows maximum activity in vitro in the pH range of enzymes in plasma?

pH range from 7 to 8

What are the activity values observed in pH as low and high?

Low as 1.5 (pepsin)High as 10.5 (ALP)

What can be controlled by the use of buffer solutions?

Pronounced effects of pH on enzyme reactions.

When does enzymes inactivation and denaturation occurs?

Occurs at temperature of 60 degrees Celsius to 70 degrees Celsius.

What is the accepted measuring temperature for enzymes in the clinical laboratory?

37 degrees Celsius

What is a substance that reduces the rate of an enzymatic reaction and is classified as reversible or irreversible?

Inhibitor

What is a structural analog of the substrate that combines with the free enzyme and competes with the normal substrate for binding at the active site?

Competitive inhibitor

What is structurally different from the substrate, it binds at a site on the enzyme molecule that is different from the substrate-binding site?

Noncompetitive inhibitor

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What is the small molecule or ion that increases the rate of an enzyme-catalyzed reaction by promoting formation of the most active state of the enzyme or of other reactants such as the substrate?

Activator

When used as analytical reagents for measuring metabolites, they increase the specificity of the substance being determined

Enzymes

What is a Coenzyme A small molecule (some contain structures derived from vitamins) that is required for some enzyme-catalyzed reactions to occur; is temporarily or permanently bound to the enzyme

NAD and NADH are examples of? CoenzymesWhat is a Prosthetic group a tightly bound, non-peptide structure required

for the activity of an enzymeWhat is a poenzyme The protein component of an enzymeH What is a oloenzyme An active enzyme formed by combination of a

coenzyme and an apoenzymeThe rate of an enzyme catalyzed-reaction is directly proportional to the amount of _____ present in the system

active enzyme

Fixed-time and continuous-monitoring methods are used to measure ______.

reaction rates

Progress of an enzyme reaction is monitored by measuring in time the decreasing concentration of the ______ or the increasing concentration of the _____.

substrate, product

The amount of substrate transformed into products during an enzyme-catalyzed reaction can be measured by?

SpectrophotometryFluorometryChemiluminescence

Immunoassays are used to measure ______instead of _______.

protein mass, catalytic activity

What assay is capable of identifying all molecules with the antigenic determinants necessary for recognition by the antibody and has been found to be significant in the determination of some digestive enzymes, such as trypsin, when inactive precursors and inhibitors of catalytic activity are present in plasma

Immunoassay

When enzymes are used as analytical reagents for the measurement of metabolites there is a great increase of _____ for the substance being determined.

Specificity

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Enzymatic methods that allow all substrate present in a reaction mixture to be completely converted into a measurable product before measurement, are referred to as

Equilibrium methods

Enzymatic methods which demonstrate that the change in substrate concentration over a fixed time interval is directly proportional to its initial concentration

Kinetic Methods

What is an immunoassay? An assay based on the reaction of an antigen with an antibody specific for the reaction.

What is antigen? 1) Any material capable of reacting with an antibody without necessarily being capable of inducing antibody formation.

2) Analyte being detected by the immunoassay.

What are antibodies? An immunoglobulin (Ig) class of molecule (IgA, IgG, IgM) that bins specifically to an antigen or hapten

Which immunoglobulin is the most widely used reagent?

Immunoglobulin G (IgG)

What is an Immunogen? A protein of a substance coupled to a carrier cable of introducing an immune response.

What is a hapten? A chemically defined determinant that by itself will not stimulate an immune response. It is an incomplete antigen

What is affinity? 1) It refers to the thermodynamic quantity defining the energy of interaction of a single antibody-combining site and its corresponding epitope on the antigen.

2) It is a property of the substance bound (antigen)

What is avidity? 1) Refers to the overall strength of the binding of antibody and antigen and includes the sum of the binding affinities of all individual combining sites on the antibody.

It is a property of the binder (antibody)What are factors affecting antigen-antibody binding?

1) Binding forces2) Reaction mechanism3) Precipitin reaction4) Chemical factors:

1)Ion species and ionic strength effects 2) Polymer effect

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In the binding forces, what are 3 major contributing forces?

1) Electrostatic van der Waals-London dipole-dipole interactions

2) Hydrophobic interactions3) Ionic coulombic bonding

In the reaction mechanism, what are the 3 phases?

1) Phase 1:Initial reaction of a multivalent antigen and a bivalent anitbody

2) Phase 2: Subsequent growth of the complexes

3) Phase 3: Precipitation of the complex before after a critical size is reached.

These soluble complexes have all antigenic sites that are covered with antibody, and lattice formation is inhibited.

Antibody Excess

This insoluble complexes (optimal proportion) state occurs when 2 to 3 antibody molecules are present for each antigen molecule; produces maximum lattice formation and therefore maximum precipitate.

Equivalence Zone

These soluble complexes have all antigenic sites that are saturated by antigen. Triplets (2 antigen + 1 antibody) are maximum size attained by particles. no precipitate is formed.

Antigen Excess

Anion and cationic salts can affect the binding of antibodies by what 2 chemical factors?

1. Ion species2. Strength effects

What are 3 examples of polymer effect in terms of having high molecular weight, high degree of linearity and aqueous solubility that can significantly increase the rate of immune complex growth and enhances precipitation of immune complex?

1. Dextran2. Polyvinyl alcohol3. polyethylene glycol

What are 2 types of qualitative methods regarding passive gel diffusion that use semisolid medium (e.g., agar agarose), and that the initial concentration of antigen and antibody are critical?

Raidal immunodifussion (RID) (single immunodiffusion) and Ouchterlony technique (double diffusion)

The term for an analytical technique that combines separations of antigens by electrophoresis with immunodiffusion against an antiserum?

Immunoelectrophersis (IEP)

Also known as two-dimensional immunoelectrophoresis, what is the term describing to drive the antigen into a gel containing antibodies specific for the antigens of interest.

Crossed immunoelectrophoresis

This technique involves an electrophoresis step, followed by transfer of the separated proteins onto on overlying strip of nitrocellulose or nylon membrane by a process called?

Electro-blotting.

This electrophoresis step involves DNA blotting onto a membrane.

Western/Southern blotting

This electrophoresis step involves RNA blotting onto a membrane.

Northern RNA

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What does RID stand for? Radial ImmunodiffusionWhat is the definition for RID? A passive diffusion method which a

concentration gradient is used for a single reactant

During RID, what happens when antibody is added?

A well defined ring of precipitation around the well indicates presence of antigen

What is also known as the “rocket” technique? ElectroimmunoassayWhat is the definition of electroimmunoassay? A type of immunoassay where a single

concentration gradient is established for the antigen and an applied voltage is used to drive the antigen from the application well into a homogenous suspension of antibody in the gel

Turbidimetric and Nephelometric assays are convenient techniques to measure what?

The rate of formation of immune complexes in vitro using their light scattering abilities

What are some examples of radioactive and non-isotopic labels?

Competitive, Noncompetitive, and heterogeneous immunoassays

What is a Heterogeneous immunoassay? Immunochemical assay that require separation from the bound label

What does RIA stand for? RadioimmunoassayDefine RIA Used radioactive isotopes of iodine, 125I and

131I and tritium (3H) as labelsWhat does EIA stand for? Enzyme ImmunoassayWhat does ALP stand for? Alkaline phosphataseWhat does HRP stand for? Horseradish peroxidaseGive some examples of enzymes that are used in EIA?

ALP, HRP, Glucose-6-dehydrogenase, B-galactosidase

What are 3 Examples of Enzyme Immunoassay (EIA)?

Enzyme-Linked Immunosorbent Assay (ELISA)Enzyme Multiplied Immunoassay Technique (EMIT)Clone Enzyme Donor Immunoassay (CEDIA)

What does Fluoroimmunoassay (FIA) use? Uses a fluorescent moleculeWhat is fluorescent used as in FIA? as an indicator label to detect and quantify

immunological reactionsWhat is Chemiluminescent immunoassay? a chemiluminescent molecule, such as

acridiniumWhat is Chemiluminescent immunoassay used for?

is used as an indicator label to detect and quantify immunological reaction

What is Electrochemiluminescent immunoassay?

an elecrochemiluminescent molecule, such as ruthenium

What is electrochemiluminescent immunoassay used for?

is used as an indicator label to detect and quantify immunological reaction

What is one definition of automation? The process whereby an analytical instrument performs many tests with only minimal involvement of an analyst

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What is another definition of automation? controlled operation of an apparatus, process, or system by mechanical or electronic devices without human intervention

What is Random-access analysis? analyses are performed on a collection of specimens sequentially and each specimen is analyzed for a different selection of tests.

What is Continuous-flow analysis? it specimen in a batch passes through the same continuous stream at the same rate and is subjected to the same analytical reactions

What is Single-channel analysis? each specimen is subjected to a single process so that only results for a single analyte are produced.

What is Multiple-channel analysis? each specimen is subjected to multiple analytical processes so that a set of test results is obtained on a single specimen.

What is multiple-channel analysis similar to? Similar to random access analysisWhat is Parallel analysis? all specimens are subjected to a series of

analytical processes at the same time and in a parallel fashion

What is Discrete analysis? the sample is aspirated into the sample probe and then is delivered, often with reagent, through the same orifice into a reaction cup or another container.

How many types of the specimen identifiers? These are: specimen labels, serial number, part number, manufacturer, and assigned patient number

What are the technologies used for automatic identification and data collection?

There are technologies used for automatic identification and data collection as barcoding, optical character recognition, magnetic stripe and magnetic ink character recognition, voice identification, radiofrequency identification touchscreen, light pen, handprint tablets, optical mark reader, smartcards.

What are the delays in preparation of specimen?

-Clotting of blood in specimen collection tubes-Subsequent centrifugation-Preparation of serum and plasma-Transfer of serum the secondary tubes

What are the methods of delivery specimen? Courier service, pneumatic tube systems, electric track vehicles, and mobile robots.

How reliable is a courier service? Courier service is usually reliable but costly and may require batches, but risk of specimen breakage or loss

How does pneumatic tube system transport? Pneumatic tube systems provides rapid specimen transportation, risk of misrouting or hemolysis

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How does specimen delivery as an electric track vehicle?

Larger capacity than pneumatic tube systems makes use of dry ice or refrigerator packs possible

How does specimen delivery as a mobile robot?

Easily adapted and programmable, but need for batching and personnel to load robots.

Once collected into a red top tube what is the first thing one must do before specimen preparation for routine tests can begin?

Red top tubes have no anticoagulant and therefore must be allowed to clot before preparing them for routine tests.

Once a red top tube is allowed to clot what is the next step in specimen preparation for routine testing?

Centrifuge the specimen for at 1200g for 10 minutes

Once a red top tube has been allowed to clot and centrifuged until separated what is the next step in preparing the specimen for routine testing?

then separate serum from the cells, labeling the secondary tube appropriately.

What are two common ways to get specimens from site of collection to the lab?

Currier service and Pneumatic tube system

What are the pros and cons of currier service. Pros-•Reliable•best way to get specimens to the lab if from a site across cities or even states.Cons-•costly•require multiple specimen batch collections and deliveries throughout the day•risk of specimen breakage•risk of specimens becoming too warm or cold if left outside in collection box.

What are the pros and cons of the pneumatic tube system?

Pros-•Rapid specimen transportCons-•Risk of specimens being sent to the wrong place and becoming lost•tube traffic may occur causing tubes to collide possibly shutting the system down -this could cause hemolysis or leaking specimens

What is an electric track vehicle? A form of specimen delivery similar to the pneumatic tube system in which specimens are carried on a track, not within a tube system, and in a specimen carrier box which can be larger and more versatile in the type of specimen being transported. Also can utilize ice packs or dry ice to keep specimens under the correct temperature conditions.

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Mobile robots are being utilized for what laboratory service?

Mobile robots are being used for specimen transport within hospitals. They are loaded with a schematic blue print of the hospital and can be programmed to go from area to area to collect and transport specimens to the proper lab.

Having instrumentation that is capable of sampling directly from a primary tube or centrifuging and aliquoting the plasma or serum into a secondary tube on its own will reduce what and increase what?

Reduce-Specimen evaporation, turn around time, risk of disease transmission, specimen contamination

Increase-efficiencyThree processes of removing protein and other interferents are?

DialysisColumn chromatographyFiltration

In immunochemistry, instrumentation is capable of separating bound and free fractions via which methods?

•coated beads•coated tubes•microtiter plates•magnetic and nonmagnetic microparticles

Define carryover in regards to specimen transport during analytical testing

The transport of a quantity of analyte or reagent from one specimen reaction into and contaminating a subsequent one.

What sample introduction system utilizes a peristaltic pump?

Continuous-flow system

What sample introduction system utilizes pipets and is highly automated.

Discrete systems

Instrumentation must have what capability to store reagents?

Refrigeration

Instrumentation must have this to identify patients and tests to be performed on each sample.

Bar code scanner

Rate of transport of reaction mixture through the system to the measurement station affects what?

Timing of the reaction

on-time events on reagent addition (or activation) relative to measurement will affect what?

Timing of reaction

What are the five main means of mixing reactants in instrumentation?

Forceful dispensingmagnetic stirringvigorous lateral displacementa rotating paddlethe use of ultrasonic energy

Some temperature control and efficient heat transfer methods seen in instrumentation are…

Air bathswater bathscontact with warm plates

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Analyzers can employ different types of optical measurement devices, list five.

PhotometersSpectrophotometersReflectance photometersFluorometersLuminometers

One way computers have impacted the clinical laboratory is:The computer controls the electromechanical operation of the analyzer. This means?

all functions are performed uniformly, in a repeatable manner and in the correct sequence

Having the computer control the automated equipment, calculation of results and monitoring of operation will increase the ___ of results.

reproducibility

One way computers have impacted the clinical laboratory is:Improved acquisition, processing and storage of operational data from the analyzers. This means?

Computers can monitor instrument functions for correct execution and record the sites and nature of a malfunction so the tech can troubleshoot and repair the problem.

One way computers have impacted the clinical laboratory is:Computer interfacing with the instrument operator. Explain this.

The instrument operator is alerted to changes in analyzer by the computer directly (ie problems, errors, low reagents, etc) so the tech can troubleshoot the problem, if needed, and the issue can be fixed.

One way computers have impacted the clinical laboratory is:Computer facilitation of communication with mainframe computer. Explain this.

This allows for LIS and instrument to be linked so results can be transferred directly to the patients records.

One way computers have impacted the clinical laboratory is:Computer integration of the monitoring functions of one or more analyzers with a single workstationExplain this?

This allows the tech to work on multiple analyzers simultaneously

Instrument cluster When several different instruments are set up to be controlled by one tech.

Work cell A robotic specimen processor monitored by a tech.

The work cell performs what tasks? Centrifuge, aliquot, label, transport and store specimens

Optical measurement devices includes: - Photometers- Spectrophotometers- Reflectance photometers- Fluorometers- Luminometers

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Impact of integration of computers into automated analyzers and analytical systems are:

- Computer control of the electromechanical operation of the analyzer

- Computer acquisition, processing and storage of operational data from the analyzers

- Computer interfacing with the instrument operator

- Computer facilitation of communication with mainframe computers

- Computer integration of the monitoring functions of one or more analyzers with a single workstation

Features of advanced analytical systems include:

• Workstations• Equipped for a defined task• Manual specimen transport• Built-in diagnostics

• Instrument clusters• Control of several instruments

by one technologist• To reduce labor costs

Work Cells consist of: - Robotic specimen preparation (Centrifuge, aliquot, label, transport and store specimens)

- Technologist as monitorAutomated Specimen Transport include: - Conveyor belts

- Robot armsAutomated Speciment Processing include: - Stand-alone specimen-processing systems

- Integrated and modular automation system- Role of process control software- Workload- 13 components of integrated systems

Name the three approaches that have been used to automatically sort specimen?

1) A conveyor belt,2) automated sorter using racks and3) stand-alone sorters.

Name the automated specimen modules which store specimens refrigerated in specific locations that are logged into a database and Adaptation of other industries’ systems to the laboratory?

Automated specimen storage and retrieval

How many types of conveyor sorting system have been used?

Three types

What is advantage of conveyor system in the first approach?

Direct sampling from a conveyor track in a loop configuration eliminates the need for separation equipment to sort specimens.

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What is disadvantage of conveyor system in the first approach?

Direct sampling may limit the rate of specimen movement on the track to the sampling speed of the slowest workstation.

What is the second approach of automated processing conveyor system?

It sorts specimen into group according to their destination in the laboratory such as for hematology or chemistry.

What is the third approach of automated processing conveyor system?

The sorter is integral to the conveyor system, and specimens are sorted as they are transported.

What are Practical Considerations that influence a laboratory’s decision to automate part or all of its operations?

Evaluation of requirements and,Problems of integration

List evaluation of requirement in practical considerations ?

Must be first step in decision to automateMapping of workflow (Box 16-4)80% rule of thumbVisits with vendors

List Problems of integration in practical considerations?

CLSI standard AUTO03-AFunctional control modelDevice integrationIntegration of process controllers and software

What is Point-of-Care Testing (POCT)? It is a method of clinical laboratory testing in which the analysis is performed at the site close to where the healthcare is provided to the patient.

What are other terms for POCT? Other terms used to describe POCT included: bedside, near patient, physician's office, extra laboratory, decentralized, off-site, ancillary, alternative site and unit-use testing.

In what types of environments would POCT be employed?

POCT would be encountered at home, community pharmacy, retail clinics, and more

What are some advantages to POCT? Reduced turnaround time, improved patient management, reduction in process error and reduction in time delays

What are the main objectives when designing a Point-of-Care instrument?

The main objectives of the design are to:1. Enable the required reaction to take place that facilitates recognition of the analyte of interest2. Ensure reliable performance of the device over a specified period of time3. Minimize the risk of error associated with the use of the device

What are key components of POCT device design?

Operator interface, bar code identification systems, sample and reagent delivery mechanisms, reaction cell, sensors, control and communications systems, data management and storage

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What are the POCT devices classified as? In vitro, in vivo, ex vivo, minimally invasiveDipsticks, complex strips, and immunostrips are examples of what type of in vitro POCT devices?

Single-use qualitative strip or cartridge and/or strip devices

What are some single-use quantitative cartridge and strip tests with a monitoring devices?

Glucose monitoring device, cardiac markers, allergy tests, fertility tests, drugs of abuse

What are the analytical principles of in vivo POCT technology?

Optical fluorescence and electrochemistry

What are the analytical principles of ex vivo POCT technology?

Optical fluorescence and electrochemistry

What are the analytical principles of noninvasive POCT technology?

Electrochemistry/ iontophoresis, andmulti wavelength spectrophotometry

What are the analytes used in in vivo POCT technology?

pH, blood gases, subcutaneous glucose

What are the analytes used in ex vivo POCT technology?

pH, blood gases, electrolytes, glucose

What are the analytes used in noninvasive POCT technology?

Transcutaneous glucose, bilirubin

How to POCT devices interface with laboratory information systems?

Electronic transfer of data from the analyzers to the LIS and into the patient's electronic medical record

What does CIC stand for? Connectivity Industry ConsortiumWhat is CIC? Developed point-of-care communication

standards which facilitate linking of devices to information management system

What are POCT01-A2 user requirements? CLSI connectivity standards to ensure that the POCT devices meet critical user requirement

What is the formula for an amino group? NH2What is the formula for a carboxyl group? COOHWhat is an amino acid? Organic compounds containing both amino

and carboxyl functional groupsPeptides and proteins are made of which basic structural unit?

Amino acids

What is the definition for ampholyte? A zero net charge as the negative and positive charge balance at a neutral pH

What is another term for ampholyte? ZwitterionAmino acid metabolism is synthesized from what?

Alpha-keto acids

What is the daily requirement for protein intake for adults?

About 0.8 g/kg body weight

What is the primary source of amino acids for protein synthesis?

Dietary protein

What is another source of free amino acids? Endogenous proteinWhat happens when the is an absence of protein uptake?

Breakdown of muscle protein serves as a source of amino acids

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What are essential amino acids? Amino acids that are not synthesized by humans and are essential dietary constituents for maintaining health or growth

Where are essential amino acids usually obtained from?

Meat, milk, eggs, and fish

How many essential amino acids are there? 8Name all the essential amino acids Isoleucine, leucine, lysine, methionine,

phenylalanine, threonine, tryptophan, and valine

What is Kwashiorkor? A diet with adequate calories with low protein intake

How is Kwashiorkor obtained? Through malnutrition due to protein deficiency

How is Kwashiorkor characterized? Decreased serum albumin, edema, ascites, growth failure, immune deficiency, and apathy

Deficiency of both calories and protein is known as?

Marasmus

What causes marasmus? Protein-calories malnutritionWhat is caused by marasmus? Generalized muscle wastingWhat is the clinical implication known as primary aminoaciduria?

A disease due to an inherited enzyme defect

Where can the defect be located? In the pathway which a specific amino acid is metabolized or in a transport system for an amino acid

What is secondary aminoaciduria? Disease of an organ such as the liver which an active site of an amino acid metabolism, or generalized renal tubular dysfunction

What is the main clinical use of amino acid analysis?

Detection and monitoring of inherited disorders

What is the most abundant amino acid? GlycineWhat type of specimens are used for the analysis of amino acids?

Heparanized plasma, urine, and CSF

When do amino acids exhibit high diurnal variations?

During mid-afternoon

When do amino acids exhibit low diurnal variations?

In the early morning

The analysis of amino acids require a normal diet for how many days before collection?

2-3 days

True or False?Blood and urine specimen should be collected simultaneously

True

True or False?All medications don’t have to be noted

False, all medications need to be noted

Why should specimen be processed rapidly and frozen rapidly?

To preserve glutamine

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Why does plasma need to be separated from whole blood rapidly?

So homocysteine can be measured accurately

How do most procedures entail removal of proteins?

By precipitation or ultrafiltration

What does TLC stand for? Thin-layer ChromatographyName 3 screening tests for the analysis of amino acids

TLC, Guthrie test, and liquid chromatography-tandem mass spectrometry

What does LC-MS/MS stand for? Liquid chromatography-tandem mass spectrometry

What does CE stand for? Capillary electrophoresisWhat does GC stand for? Gas chromatographyName quantitative tests for the analysis of amino acids

Ion exchange chromatography with post-column reaction with ninhydrin, reversed-phase high-performance liquid chromatography, LC-MS/MS, CE, and GC

What is a protein? A polymer of amino acidsHow are amino acids linked? By peptide bonds with a specific sequence

that folds into a defined structureProteins contain a group of complex organic compounds that contain which elements?

Carbon, hydrogen, oxygen, nitrogen, and usually sulfur.

Which is the characteristic element? nitrogenWhat is proteome? The total complement of proteins present at

the time in a call or cell type expressed by the genetic material of an organism

Proteome defines the filed of? proteomicsA peptide is a compound that consists of how many amino acids linked in a chain?

Two or more via peptide bonds

Define a peptide bond The amide bond that is formed between the carboxyl group of one amino acid and the amino group of another

True or FalseProteins act as structural components

True

Proteins facilitate catalysis of what kind of reaction?

Chemical reactions including synthesis of DNA, RNA and proteins

How can proteins generate energy? Through electronic transferTrue or FalseProteins can serve as ion channels and pumps

True

What kind of molecules can proteins act as? Carrier moleculesTrue or False?Proteins can effect immune defense

True

Proteins can act as what for intercellular communication?

Receptors, hormones, and cytokines

True or False?Proteins constitute signaling networks for extracellular communication

False, proteins constitute signaling networks for intercellular communication

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Proteins are covalently linked how? by peptide bondsthe α-amino group of one amino acid is link covalently bonded to what?

with the α-carboxyl of a second amino acid

What is the amino acid with the free amino group in the peptide termed?

N-terminal residue

What is the amino acid with the free carboxyl group termed?

C-terminal residue

Where does protein synthesis begin? with the N-terminal residueWhat are peptides named from? the N-terminal residueWhat are Primary structure? sequence of amino acids in a peptide or

proteinWhat is the sequence of amino acids comprising the polypeptide chain that is genetically encoded?

deoxyribonucleic acid (DNA).

The following are extra questions from 18:22-38What does the secondary structure of a protein describe?

The recurring formations along the polypeptide chain caused by intramolecular forces such as hydrogen bonding

What are the three parts of the secondary structure?

α-helix,β-pleated sheets, and random coil

What does random coil refer to? The segments of protein that lack secondary structure.

What does tertiary structure of a protein refer to?

The folding of the chain of amino acids into a three dimensional structure

How can the tertiary structure be described? Folded, three-dimensional conformation of the protein that is stabilized by both covalent bonds and non-covalent forces mainly involving amino acid R group

What has a great effect on a proteins tertiary structure?

The proteins environment

What can cause denaturation of proteins? Mechanical agitation, heat or extreme chemicaltreatment

What does quaternary structure of a protein refer to?

The association of multiple peptide bonds

Association of what forms a functional protein complex?

Two or more polypeptide chains, or subunits.

What happens to molecular size of proteins when they are denatured?

Size increases

How can proteins be separated? gel filtration chromatographygradient pore gel electrophoresisultracentrifugationpolyacrylamide gel electrophoresisMass spectrometry

What factors can affect protein solubility? pH, ionic strength, temperature, dielectric constant of the solvent.

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Is albumin soluble in water? YesWhat else is albumin soluble in? dilute and moderately concentrated salt

solution (ammonium sulfate)What is albumin insoluble in? saturated and concentrated salt solutionWhat is globulin soluble in? hydrocarbon solvents

weak salt solutionIs globulin soluble in water or saturated and concentrated salt solution?

No

What is the solubility of albuminoids? Insoluble in most common reagentsWhat enables proteins separation by electrophoresis, isoelectric focusing, and ion exchange chromatography?

The variable charge and isoelectric points

On what basis does differential adsorption separate polypeptides and other molecules?

On the basis of their affinity for a hydrophobic stationary phase

What are two examples of differential adsorption?

Liquid chromatography, and reversed-phase chromatography

What is affinity chromatography? Highly specific molecular interactions to bind a protein or a peptide to a stationary phase, while other proteins and peptides are removed

What may Affinity purification of antibodies or proteins be used as?

A preparative technique to prepare specific antibodies or proteins as assay components

What is density gradient ultracentrifugation used for?

Used to separate some protein complexes that have a different density when compared with most proteins, such as lipoproteins with low density but with high lipid content

What are the physical methods of quantitative analysis?

Dye bindingDirect absorbance measurementsMeasurement of protein ligandsSelective precipitationMass spectrometry

What are two other types of quantitative analysis?

Activity measurements and immunoassays

What are the methods of qualitative alalysis of proteins?

ElectrophoresisChromatographyGenetic analysisFunctional assaysMass spectrometry

What is the Kjeldahl method in measurement of total protein?

A standard reference method based on nitrogen determination

What is the standard reference for N2 in the Kjeldahl method?

1 gram N2 = 6.25 grams of protein

What is the principle of biuret method? Cupric ions in the alkaline copper tartrate reagent react with peptide bonds to form a blue violet complex proportional to the number of peptide bonds present.

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What does the analysis depend on in the biuret method?

The presence of peptide bonds

What is the composition of the biuret reagent? Alkaline copper tartrate as a source of cupric ionsRochelle salt (NaK tartrate) is a complexing agent to keep copper in solutionSodium hydroxide for alkalinityPotassium iodide which prevents autoreduction of copper

What is direct spectrophotometric measurement of total protein?

Measurement of protein content using absorbance of ultraviolet (UV) light at 200 to 225 nm and 270 to 290 nm.

What is the principle of the dye-binding method for measuring total protein?

Dye binds to protonated amine group of amino acids with absorption at 595 nm

What are the commonly used dyes? Pyrogallol red (for urine and CSF)Coomassie brilliant blue

For measurement of total protein, what is the method of direct spectrophotometric?

Measurement of protein content using absorbance of ultraviolet (UV) light at 200 to 225 nm and 270 to 290 nm.

How is the dye-binding method for measurement of total protein?

Dye binds protonated amine group of amino acids with absorption at 595 nm

What dyes are commonly used? Pyrogallol red (for urine and CSF)Coomassie brilliant blue

What is the Folin-Ciocalteu (Lowry) method? That is a measurement of refractive index to measure total solutes in urine specimen

What is a refractometry method for measurement of total protein?

That is a measurement of refractive index to measure total solutes in urine specimen

How are the turbidimetric and nephelometric methods based on the formation of aggregates or precipitation when every agent?

These are added to lower protein solubility or when an antibody is added to a protein

What are precipitation reagents? Trichloroacetic acid (TCA)Sulfosalicylic acid (SSA)Sulfosalicylic acid combined with sodium sulfateBenzethonium chlorideBenzalkonium salts under alkaline conditions

What is the formula to calculate for normal A/G ratio?

Normal A/G ratio = 1 – 2.5:1

What is the A/G ratio if albumin is greater than globulin?

A/G = Albumin/Globulin: 1

What is the A/G ratio if globulin is greater than albumin?

A/G = 1: Globulin/Albumin (Inverted A/G ratio)

What is Serum Protein Electrophoresis (SPE) performed at pH 8.6 in the low ionic strength buffer?

Proteins with a negative charge at pH 8.6 move toward the anode (positive pole)Proteins with a positive charge moved towards the cathode (negative pole)

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How many types of supporting media in SPE are there?

These are papers, garose gel, cellulose acetate, starch gel and acrylamide gel

What are stains used in serum protein electrophoresis?

Amido black and ponceau SCoomassie brilliant blue

What are silver stains used for? Silver stains are used for nanogram concentrations of proteins and nucleic acid, they are more sensitive than Coomassie blue stainThey are also used for CSF and urine proteins, shades of green, yellow, blue and red

What is the intensity of staining individual bands of protein generally?

It is proportional to the amount of protein in a band

What is a quantitative assessment in SPE done?

Quantitative assessment is done by densitometry of stained hands

In electrphoresis, when the protein is too high a concentration, what is each band appearance?

Each band in the stained pattern should be uniformly colored; holes or doughnut-like appearance within a band occurs

How is the order of migration in SPE? The order is following Albumin, Alpha1-globulin, Alpha2-globulin, Beta-globulin, Gamma globulin

What is a serum preferred? Serum is the preferred specimenWhere extra fraction will show if plasma is used?

It will show in the beta-gamma region, and fibrinogen is a beta globulin

What is the main application of SPE? The main current application of SPE is for the detection or quantification of monoclonal immunoglobulins that occur in disorders, such as multiple myeloma.

How does monoclonal immunoglobulins usually appear?

Monoclonal immunoglobulins usually appear as sharp bands in the gamma (γ) or beta (β) region

How different between monoclonal immunoglobulins and normal immunoglobulin’s?

Monoclonal immunoglobulins differ from normal immunoglobulins, which migrate as a diffuse zone, mainly in the gamma region

What are 5 types of supporting media for serum protein electrophoresis (SPE)?

PaperAgarose gelCellulose acetateStarch gelAcrylamide gel

3 Stains used for SPE’s include ? Amido blackPonceau SCoomassie brilliant blue

What silver stain used for? - nanogram concentrations of proteins and nucleic acid- more sensitive than Coomassie blue stain- used for CSF and urine proteins- shades of green, yellow, blue and red

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Serum Protein Electrophoresis (SPE) -the intensity of staining individual bands of protein generally is proportional to the amount of protein in a band-quantitative assessment is done by densitometry of stained hands-In electrophoresis, each band in the stained pattern should be uniformly colored; holes or doughnut-like appearance within a band occurs when the protein is too high a concentration.Remedy : Dilute elevated specimens before re-running the electrophoresis

What is the order of migration? 1.Albumin2.Alpha1-globulin3.Alpha2-globulin4.Beta-globulin5.Gamma globulin

What if plasma is used as a specimen? - extra fraction will show in the beta-gamma region- Fibrinogen is a beta globulin

What is the preferred specimen in protein electrophoresis?

Serum

What is the main current application of SPE? is for the detection or quantification of monoclonal immunoglobulins that occur in disorders, such as multiple myeloma.

What do Monoclonal immunoglobulins usually appear as?

sharp bands in the gamma(γ) or beta(β) region

What do Monoclonal immunoglobulins differ from normal immunoglobulins?

normal immunoglobulins, which migrate as a diffuse zone, mainly in the gamma region

In regards to electrophoresis, the abbreviation IFE stands for?

Immunofixation Electrophoresis (IFE)

With the application of specific antibodies, IFE can be directed against which immunoglobins/ light chains?

Immunoglobins: IgG, IgM, IgAAlso kappa (κ) and lambda (λ) light chains

IFE is used on what type of gel? Agarose gelElectrophoretic separation of urinary proteins is used to detect what?

The presence of free monoclonal immunoglobulin light chains or other low-molecular-mass proteins

What is another term for the presence of free monoclonal immunoglobulin light chains?

Bence-Jones Proteins

The detection of Bence-Jones proteins can be indicative of what disease?

Multiple Myeloma

Fill in the Blank: relies on the separation of ions in an electrical field in a vacuum?

Mass Spectrometry

In mass spectrometry, separation of molecules is purely based on what?

Mass-to-charge (m/z) ratio

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In what case would a mass spec. be ideal? Ideal for highly precise qualitative analysis of peptides

What is an Acute Phase Response (APR)? A systemic response to infection, tissue injury or inflammatory processes resulting in fever, increased WBC count, and changes in the concentration of many plasma proteins.

Proteins affected or increased by APR are known as?

Positive acute phase proteins (APPs)

Known Positive APPs include: α1-antitrypsin Fibrinogen α1- acid glycoprotein C-reactive protein Haptoglobin Serum amyloid Ceruloplasmin C4, C3

Proteins not affected or decreased by APR are called?

Negative acute phase proteins (APPs)

Known Negative APPs include: Transthyretin Albumin Transferrin

What is Prealbumin (Transthyretin)? A transport protein of thyroid hormones and RBP

Prealbumin/ Transthyretin is composed of four identical subunits that associate to form what?

A hollow core containing T3 and T4 binding sites

Binding of one hormone molecule decreases the binding affinity of the second so that only one site is normally occupied. This is known as what?

Negative cooperativity

Where is prealbumin/ transthyretin synthesized?

In the liver and to a lesser extent in the choroid plexus of the central nervous system, accounting for the relatively high concentration in CSF

Synthesis of prealbumin/ transthyretin is stimulated by what hormones and drugs?

Synthesis is stimulated by glucocorticosteroid hormones, androgens and many nonsteroidal anti-inflammatory drugs (NSAIDs) including aspirin

Prealbumin/ Transthyretin is often used as indicator of?

Protein Nutrition

Prealbumin/ Transthyretin is increased during what conditions?

With corticosteroid or NSAID therapyHodgkin Disease

Prealbumin/ Transthyretin is decreased during what conditions?

Inflammation and malignancy Cirrhosis of the liver Protein-losing diseases of the gut or

kidneysWhat is the migration pattern for prealbumin/ Migrates as a minor band anodal to albumin

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transthyretin on Serum Protein Electrophoresis (SPE)?Name 2 methods that can measure prealbumin: Immunoturbidimetric or

immunonephelometric methodsWhat is Retinol-Binding Protein (RBP)? A transport protein bound to prealbuminWhat element is required for the synthesis of RBP?

Zinc

What is required for RBP to transport out of the Golgi apparatus?

Retinol

Where is RBP absorbed in the body? It is absorbed by the proximal renal tubular cells and catabolized

Fill in the Blank:Serum concentration of RBP is in chronic renal disease.

Increased

RBP is increased with? Corticosteroid NSAID therapy And in Hodgkin Disease

RBP is decreased in? Liver disease, protein malnutrition, and acute phase response (APR)

What is the most abundant protein in the plasma from mid-gestation until death?

Albumin

Albumin is a major protein component of most extra vascular body fluids including?

CSF Interstitial fluid Urine Amniotic fluid

Albumin is synthesized by? The hepatic parenchymal cellsAlbumin is a major transporter of? Free fatty acids, bilirubin, calcium, thyroid and

steroid hormones, drugs and thiol-containing compounds

Albumin is increased during? Dehydration Prolonged application of the tourniquet Specimen evaporation before analysis

Albumin is decreased in? Analbumemia – a rare genetic deficiency with plasma albumin concentration less than 0.5 g/L

Inflammation – Albumin is a negative APP. Acute and chronic inflammation are common causes of hypoalbuminemia

Hepatic disease

How does hepatic disease decrease the concentration of albumin?

Loss of parenchymal cells function by 50%

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Nutritional deficiency Increased distribution into the

extravascular space Direct inhibition of synthesis by toxins

such as alcoholHow is albumin decreased in urinary loss? - Small increases in albumin excretion to

>30 mg/d indicate early stages of glomerular or tubular injury and risk of progression to more severe kidney disease (Microalbuminuria)

- Severe glomerular injury produces the nephrotic syndrome, which is characterized by excretion of >3.5 g/d of protein (mainly albumin).

How is albumin decreased in Gastrointestinal loss?

Inflammatory disease of the intestinal tract is associated with increase gastrointestinal loss of albumin

How is albumin decreased in protein energy malnutrition (Marasmus)?

Albumin concentration help to detect and monitor protein nutritional status

How is albumin decreased in burn injury? Burn patients experience severe loss of albumin from wounds due to the combined effects of epithelial losses, accelerated catabolism and the APR

How is albumin decreased in Edema and ascites?

Edema and ascites are usually secondary to increased vascular permeability, and albumin levels are decreased as a result of redistribution of albumin into extravascular space

Whsat are two ways for Albumin Determination?

- Dye-Binding methods• Bromcresol green (BCG)• Bromphenol blue (BPB)• Bromcresol purple (BCP)• Hydroxyazobenzenebenzoic acid

(HABA)- Immunoturbidimetric and

nephelometryWhat is High-Density Lipoprotein? Also called alpha lipoprotein or good cholesterol

Characterized by high density because of its protein and phospholipid contentApolipoprotein A1 the major protein, is a negative APP that is considered to have anti-inflammatory properties

What is α1-Acid Glycoprotein? - Also known as orosomucoid- One of the highest concentration

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proteins in the alpha-1 globulin religion- Amember of the lipocalin family of

proteins that bind lipophilic substances, such as progesterone and basic drugs including propanolol, chlorpromazine, quinidine, cocaine and benzodiazepine.

- It is synthesized mainly by hepatic parenchymal cells

- A positive APPWhat factors does α1-Acid Glycoprotein increases in?

- G.I. inflammatory disease- Malignant neoplasms- Corticosteroids and NSAIDs

medicationWhat factors does α1-Acid Glycoprotein decreases in?

- Estrogen medication- Protein-losing syndrome such as

nephrotic syndromeHow is electrophoresis used for α1-Acid Glycoprotein Determination?

Using periodic acid Schiff or other carbohydrate stains

How is α1-Acid Glycoprotein Determination quantified?

By immunochemical methods, including turbidimetry and nephelometric.

What is α1-Antitrypsin (AAT)? - Is a serpin (serine proteinase inhibitor) that inactivates serine proteases such as neutrophil elastase

- Synthesize primarily by hepatic parenchymal cells

- It is the most important inhibitor of leukocyte elastase, as release by neutrophils

- Levels are elevated by the APR and by estrogen oral contraception

- Levels are low in individuals with neonatal respiratory distress syndrome, severe pancreatitis and protein-losing disorders

- Quantified by immunoturbidimetry or immunonephelometry

- Phenotyping of AAT is performed by isoelectric focusing

What is α -Fetoprotein (AFP)? - Is the most abundant plasma proteins in the early embryonic life

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- Homologous to albumin but differs in having about 4% carbohydrate

- Serum concentration serves as a tumor marker for Hepatocellular and germ cell carcinoma

- Maternal serum concentrations are analyzed as indicators of fetal trisomy of chromosome 18 or 21

What is Haptoglobin (Hp)? - Concentrations are increased by glucocorticosteroids, androgens and many NSAIDs

- Levels are elevated in selective protein-losing syndrome such Nephrotic syndrome

- Significantly elevated in biliary obstruction in the absence of severe hepatocellular disease

- Measured by immunoturbidimetry and immunonephelometry

What is α2-Macroglobulin (AMG)? - A major plasma proteinase inhibitor- A very large molecule that does not

diffuse from the plasma space into extracellular fluids in significant amounts

- Homologous to complement components C3 and C4 in inhibiting classes of proteinases, including those with serine, cysteine and metal ions in their catalytic sites

- Synthesized by the liver- Levels are higher in women of

childbearing age and in infants and children

- Increased in nephrotic syndrome- Decreased in severe acute pancreatitis

or with advance carcinoma of the prostate

- Ccan be measured by zone electrophoresis in the alpha2-globulin region

- Quantified by immunoturbidimetry or immunonephelometry

What is Ceruloplasmin (Cp)? - An α2-globulin that contains approximately 95% of the total serum

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copper, giving it a blue color- May give plasma a greenish tint when

concentrations are significantly elevated during pregnancy

- Synthesized primarily by the liver, and is responsible for catalysis of reduction and oxidation reactions

- Important in regulating the ionic state of iron, oxidizing from the ferrous (Fe2+) to the ferric (FE3+) state, thus permitting incorporation of the iron into transferrin

- It is a weak, late-reacting positive APP- Increased significantly by estrogens, as

in pregnancy or with the use of oral contraceptives

Following are clinical significant of what type of plasma protein?decreased levels are due to dietary copper insufficiency (including malabsorption), associated with neutropenia, thrombocytopenia, low serum iron and hypochromic, normocytic, or microcytic anemia that does not to respond to iron therapy

Ceruloplasmin (Cp)

What disease is characterized by increased total body copper and deposited in tissues including the hepatic parenchymal cells, the brain, and the periphery of the iris (resulting in the characteristic Kayser-Fleisher ring)?

Wilson disease, or hepatolenticular degeneration

How measure Ceruloplasmin (Cp)? by immunoturbidimetry and immunonephelometry

What is separation and transportation or holding condition for Ceruloplasmin (Cp)?

serum or plasma should be separated from blood as soon as possiblespecimen should be refrigerated for up to three days or frozen at below -70°C for prolonged storage

What is concentration of Ceruloplasmin (Cp) at birth?

Low at birth

When is peak concentration of Ceruloplasmin (Cp)?

increased to peak concentrations at 2 or 3 years of age

What is concentration of Ceruloplasmin (Cp) for adult?

decline slowly until adolescence, when adult concentrations are reached

What is other name for Transferrin? also known as siderophilin

What is the major plasma transport protein for iron?

Transferrin

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What is the main factor in plasma which Transferrin accounts for it?

total iron binding capacity

Where is Transferrin in the body synthesized and how it migrates in electrophoresis?

synthesized primarily in the liver and usually migrates in the β1-region on routine clinical electrophoresis of serum

What is relationship between Transferrin and iron concentration?

plasma concentrations are inversely related to iron concentration

What is concentration of Transferrin and iron saturation in hereditary hemochromatosis (iron-overload state)?

concentration is normal in hereditary hemochromatosis (iron-overload state), but iron saturation often exceeds 55%

What is diurnal cycle for Transferrin? serum iron concentration usually is highest in the morning and decreases markedly in the evening

What is level of Transferrin in pregnancy and inflammation?

high concentrations are present in pregnancy and during estrogen administrationis a negative APP, and low concentrations occur in inflammation and malignancy

What is the level of synthesis of Transferrin in following diseases?chronic liver disease, protein malnutrition, protein loss as seen in nephrotic syndrome or in protein-losing enteropathy

Decreases

What are laboratory considerations for Transferrin determination?

Serum electrophoresisImmunoturbidimetry and Immunonephelometry methodsPhotometric assays for TIBC (Total Iron Binding Capacity)

What is the formula for TIBC (Total Iron Binding Capacity)?

TIBC = UIBC + IronWhere: TIBC = total iron binding capacity UIBC = unsaturated binding capacity

Transferrin (TRF) migrates in…………on routine serum electrophoresis.

β1 –region

What is equation which TIBC is used to estimate TRF?

TRF (mg/dL) = 0.70 x TIBC (µg/dL) or

Iron% Transferrin saturation = --------------- x 100

TIBCWhat Low-Density Lipoprotein is termed on the basis of its electrophoretic mobility?

β-lipoprotein

What percentage of LDL is lipid? 80%What percentage of LDL is protein? 20%What is the main lipid component of LDL? cholesterol esterWhat is the main protein component of LDL? apolipoprotein BIncrease in LDL concentration confer increased risk of ……………..?

cardiovascular disease

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How concentration of LDL are assessed? assessed by calculation of LDL cholesterol that are based on other lipid measurements, direct quantification of LDL cholesterol, or quantification of apolipoprotein B

Following is characteristic of what plasma protein?a low-molecular-mass protein found on the cell surfaces of all nucleated cells

β2-Microglobulin (BMG)

Which protein plasma is the light or beta chain of the human leukocyte antigens (HLAs)?


Which protein is high in individuals with renal failure, inflammation, and neoplasm especially those associated with B lymphocytes


Name a protein marker for renal tubular function?

urinary BMG (β2-Microglobulin)

BMG levels has been used as a staging criterion for what type of disease?

multiple myeloma

Name a protein plasma which can deposit as amyloid, leading to systemic amyloidosis in many patients of long-term dialysis?


What protein is the terminal component of the coagulation system, aggregates to form a fibrous network when it is cleaved by the protease thrombin?

Fibrinogen

What protein is a highly elongated protein that consist of six polypeptide chains joined by disulfide bonds?

Fibrinogen

Fibrinogen is positive or negative APP? PositiveWhat protein is the major contributors to plasma viscosity because of its large size?

Fibrinogen

What does occur when Fibrinogen is low? the result of consumption from extensive bleeding or dysregulation of the coagulation system

What is the level of Fibrinogen in disseminated intravascular coagulation (DIC)?

Low

In what disease Dysfibrinogenemia (functional disorders of fibrinogen) may be seen?

Liver disease

What high levels of Fibrinogen on a chronic basis are associated with?

increased risk of cardiovascular disease, which may relate to increase plasma viscosity or the increased tendency for thrombosis

One of the assay to measure Fibrinogen is? functional clotting assaysWhat method can be used to measure Fibrinogen level?

measured by immunoturbidimetry methods or by turbidimetric methods based on the addition of precipitating reagents

What protein may be mistaken for a paraprotein if not completely removed from

Fibrinogen

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serum on serum protein electrophoresis?Where C-reactive Protein (CRP) is found? in the sera of acutely ill individualsName the bacteria that CRP is able to bind the cell wall C polysaccharide of it?

Streptococcus pneumoniae

CRP is positive or negative APR? an early positive APRCRP used extensively as a marker of……….? inflammationWhere CRP synthesized in the body? LiverWhat is function of CRP? to protect the body against foreign organisms

and to assist in clearing tissue debrisCRP binds to a variety of compounds is dependent on the presence of …………?

Calcium

What is diagnostic window for CRP? Levels begin to rise within 6 to 12 hours of the onset and peaks within 48 hours

In what disorders CRP increases? myocardial infarction, stress, inflammation, trauma, neoplastic proliferation, infection, surgery

CRP is a stronger stimulus in bacteria or viral infection?

Bacterial infection is usually a stronger stimulus than viral infection

Name the protein marker for bacterial infection in newborns?

CRP

What protein marker is used to assess the activity of disorders such as rheumatoid arthritis and Crohn disease?

CRP

What are referred of High-sensitivity CRP (hsCRP) assay?

Detection limit of about 1 mg/Llevels greater than 2 mg/L is suggestive of increased cardiovascular risk

What types of assays are used for high sensitivity CRP?

Particle-enhanced immunoturbidimetry, nephelometry, or sandwich immunoassays with fluorescence or chemiluminescent detection.

What type of gel is used for electrophoresis of CRP?

Cellulose acetate or agarose gel

What are complement proteins and where are they found?

Complement proteins help to complement the ability of antibodies and phagocytic cells to clear pathogens from an organism. They are found in normal blood plasma and also as cellular receptors on cell surfaces.

What complement proteins are in the Classical Pathway?

It includes C1, C4, C2, and C3 (in order of activation)

How is the Classical Pathway activated? It is activated primarily by complexes of immunoglobulins with antigen or bacteria or other ligands with CRP

What complement proteins are in the Alternative Pathway?

Includes C3, factor B and D, and properdin

How is the Alternative Pathway activated? It is activated by bacterial lipopolysaccharides, cellular proteases and

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cobra venomWhat complement proteins are in the Lectin Pathway?

It includes mannan-binding protein (MBP), ficolins, and proteases

How is the Lectin Pathway activated? It is activated by binding of ficolins and MBP to mannose-rich oligosaccharides that are present in the cell walls of many microorganisms

What complement proteins are in the Membrane Attack complex?

Includes C5 through C9 and inserts into cell membranes and lyses cells

What protein is activated through all of the pathways?

The common step involved is the activation of C3 to C3b.

What is chemotaxis? The movement or orientation of an organism or cell along a chemical concentration gradient or in response to a chemical stimulus

What is opsonization? It refers to an immune process where particles such as bacteria are targeted for destruction by an immune cell known as a phagocyte

What is phagocytosis? The engulfing and ingestion of bacteria or other foreign bodies by phagocytes to form an internal vesicle known as a phagosome

Where are complement components mainly synthesized?

The liver

Of all the complement proteins, which is the most abundant?

C3

What does a genetic deficiency of C2 and C4 cause?

A genetic deficiency of C2 and C4 is associated with autoimmune immune complex diseases such as systemic lupus erythematosus (SLE), polymyositis and glomerulonephritis.

What causes hereditary angioedema? Heterozygous deficiency of C1 inhibitor results in hereditary angioedema (HAE)

What are coagulation proteins? A complex system designed to prevent bleeding from injuries by forming blood clots

What organ primarily produces all of the coagulation factors?

The liver

Of all the coagulation factors, which is the most important?

Fibrinogen

What causes low concentrations of clotting factors?

The result of hereditary deficiencies, consumption by active clotting, liver disease or anticoagulant therapy

What are some hereditary disorders that cause excessive bleeding?

Hemophilia A, factor VIII deficiency, hemophilia B, and factor IX deficiency

What may cause a risk of thrombosis? Risk of thrombosis may be result of factor V Leiden deficiency, antithrombin III deficiency and protein C deficiency

What are immunoglobulins (antibodies)? They’re a family of proteins the contain highly specific antigen-binding sites.

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What stimulates the production of immunoglobulins, and what do they attack?

Synthesis of immunoglobulins is stimulated by foreign immunogens, leading to immunoglobulins that selectively bind foreign molecules or organisms.

What is the structure composition of immunoglobulins?

All immunoglobulins include one or more basic units consisting of two identical heavy (H) chains encoded on chromosome 14 and two identical light (L) chains encoded on chromosome 2 that are joined by disulfide bonds.

On what chromosomes are the light and heavy chains encoded?

Heavy chain is encoded on chromosome 14 and the light chains on chromosome 2

What forms the antigen-binding site on the immunoglobulins?

Two variable domains, one from the light chain and one from the heavy chain, together form an antigen-binding site.

What are heavy chains made up of? They consist of gamma, alpha, mu, delta, and epsilon

What are light chains made up of? Kappa and LambdaWhere are immunoglobulins synthesized? Immunoglobulins are synthesized by cells of

the B-lymphocyte lineage.What are the five classes of immunoglobulins? IgG, IgA, IgM, IgE, and IgDWhich of the five immunoglobulins are the most abundant?

The most abundant class of Ig, making up 70 to 75% of total Ig. Of this amount, 65% is extravascular and 35% is found in plasma.

What does IgG do? It is responsible for long-term physiologic protection and the majority of neutralization of bacteria and virus antigens

What is the circulating half-life of IgG? Approximately 22 daysWhich, if any, of the immunoglobulins can cross the placenta blood barrier?

IgG

What are the four subclasses of Immunoglobulin G (IgG)?

IgG includes four subclasses - IgG1, IgG2, IgG3, and IgG4

- IgG1 and IgG3 strongly activate complement via the classical pathway

- IgG2 weekly activates complement- IgG4 does not activate complement- IgG1 and IgG3 bind Fc receptors on

phagocytic cells can cross the placenta via receptor-mediated active transport

IgG1 is the major neonatal Ig.

What are some characteristics of Immunoglobulin A (IgA)?

- Makes up 10% to 15% of serum immunoglobulin

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- On electrophoresis, IgA migrates near the junction of the beta and gamma regions

- IgA also activates the complement by the alternative pathway

What are two types of Immunoglobulin A (IgA)?

- Plasma dimeric IgA1: sedimentation coefficient of 7S

- Secretory dimeric IgA2: sedimentation coefficient of 11S and present in saliva, tears, sweat, milk, colostrum and secretion of nasal, gastrointestinal and tracheobronchial origin

Where is Immunoglobulin A (IgA) synthesized and secreted?

- It is synthesized mainly by plasma cells in the mucous membranes of the gut and bronchi and in the ductules of the lactating breast

- This secretory component helps transport secretory IgA across mucosal epithelium and into secretions

- Secretory IgA is more abundant than IgG in colostrum and milk and may help protect neonates from intestinal infection

What is Immunoglobulin M (IgM)? - Produced at early stages of β-cell development.

- Comprises about 5%-10% of the total circulating immunoglobulins

- Largest immunoglobulin, a pentamer (molecular weight > 900 kilodaltons)

- Most primitive immunoglobulin and is found on the surface of early B lymphocytes

- First or primary response immunoglobulin to be produced following antigen assault.

What is Immunoglobulin E (IgE)? - Monomeric immunoglobulin in plasma- Lowest concentration (about 0.05

mg/dL)- Attaches rapidly to the membranes of

mast cells by its Fc region- Cross-linking of two IgE molecules by

antigen causes release of histamine, heparin and vasoactive amines by mast cells

What role does Immunoglobulin E (IgE) play in allergic or hypersensitivity reaactions?

- Urticaria- Hay fever

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- Asthma- Eczema

What is Immunoglobulin D (IgD)? - A monomeric immunoglobulin in plasma

- Active on the cell surface of B lymphocytes

- Labile upon storage- Not routinely measured

What is the significance of immunoglobulin deficiency?

- Immunodeficiency may be the result of a deficiency of a single factor or of a combinations affecting multiple systems and factors

- It results in increased susceptibility to infections that usually do not cause disease in people with normal immune systems

What is Polyclonal Hyperimmunoglobulinemia?

- Polyclonal increases in plasma immunoglobulins constitute the normal response to infection

- IgG response predominates in most autoimmune responses; IgA in skin, gut, respiratory and renal infections; and IgM in primary viral infections and with bloodstream parasites like malaria.

What are some characteristics of Monoclonal Immunoglobulins (Paraproteins)?

- Produces a discrete band on electrophoresis often referred to as an M-spike or M-protein

- up to 25% of paraproteins are benign and have been termed monoclonal gammopathy of undetermined significance (MGUS)

- Bence-Jones proteins: the primary clinical interest in identifying paraproteins involves the detection or monitoring of proliferative disorders of B lymphocytes

What malignant neoplasm is usually a single clone of plasma cells?

Multiple myeloma

What is the tumor called when the plasma cells proliferate diffusely throughout the bone marrow?

plasmacytoma

What is produced due to multiple myeloma? Osteolytic bone lesionsWhat are decreased due to multiple myeloma? Bone marrow cells like platelets, red cells,

neutrophilsWhat is inhibited due to multiple myeloma? Normal plasma cellsWhat consequently happens? Synthesis of other immunoglobulins is

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reduced, recurrent infections may occurLymphomas or chronic lymphocytic leukemias are examples of ___.

Lymphoid tumors

Where do lymphoid tumors arise? From less mature stages in B-lymphocyte development

What is a malignant proliferation of plasmacytoid cells producing IgM?

Waldenstroms Macroglobulinemia

What causes hyperviscosity that may require treatment by plasma exchange?

IgM and IgM paraproteins

What is the rare disorder in which a free Ig heavy chain is produced?

Heavy Chain disease

What is the most common form of Heavy Chain disease?

Alpha chain disease

What is alpha chain disease associated with? The heavy chain of IgAWhat can cause malabsorption in alpha chain disease?

Intestinal infiltration with clonal lymphocytes

What are serum proteins or protein complexes that precipitate at a temperature lower than normal core body temperature?

Cryoglobulins

What is the result of precipitation in tissues? Vasculitis and ischemic injury to peripheral tissues of the lower temperature

How do you lower a patient's cryoglobulin concentrations?

Patient must be kept in a warm environment

What do type I cryoglobulins consist of? Monoclonal immunoglobulins, often IgMHow many days are serum usually refrigerated for cryoglobulin analysis?

7 days

What is the next step after 7 days? CentrifugationWhat happens when cryoglobulins that precipitated in the cold are warmed back to 37C?

They should redissolve

What is a pathological process in which proteins are deposited in tissues as aggregates (amyloid), and form b-sheet structures?

Amyloidosis

What can result from amyloidosis? Kidney failure, heart failure, and peripheral neuropathies

What are stable structures resistant to proteolysis and normal clearance mechanisms?

Aggregates

Aggregates accumulate over time. TRUEWhat happens when there is increased production of specific proteins, genetic variants of proteins, or fragments of proteins?

Formation of amyloid

Where are the tissues obtained in order to diagnose amyloidosis?

From fat aspirates, rectal biopsy specimens, or biopsy specimens of affected tissues

What can the b-sheet structure of the amyloid be stained with?

Congo red dye

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What occurs as the result of increased concentrations of proteins including......Ig light chain paraproteins?

AL amyloidosis

...BMG in renal failure? AB2 amyloidosis

...Serum amyloid A AA amyloidosisUrine usually contains much lower protein concentration than plasma.

TRUE

How does urine maintain its concentration? Through the combined action of glomerular filtration and tubular uptake of protein

Why is analysis of urinary proteins mainly performed?

To diagnose kidney disease or disorders with paraprotein production

What is the increase of urinary protein excretion called?

Proteinuria

When can cause proteinuria? Glomerular injury, postrenal proteinuria, tubular injury, overflow proteinuria of low molecular weight proteins

Why is glomerular proteinuria sometimes referred to as albuminuria?

Because albumin is the major protein

Glomerular proteinuria is seen in conditions including diabetes, systemic lupus erythemotosis, IgA nephropathy, glomerularnephritis, hepatitis and HIV infections.

TRUE

What is characterized by hypoalbuminemia, hypercholesterolemia, and edema

Nephrotic syndrome

Excretion of ____ g/d protein is a criterion for nephrotic syndrome.

>3.5

What signifies early stage of glomerular injury and a risk factor for progressionof kidney disease in diabetes?

Microalbuminuria (30-300mg/d)

How is urine albumin detected? By dipsticks, followed by quantitative assays like pyrogallol red dye binding assays and benzethonium chloride turbidity assays

When can functional or benign proteinuria occur?

With exercise fever and exposure to cold

Postural or orthostatic proteinuria is associated with the upright position.

TRUE

First voided urine is analyzed to detect orthostatic proteinuria.

TRUE

What is the appearance of low molecular weight proteins in the urine as a result of decreased reabsorption by the proximal tubules?

Tubular proteinuria

What is Fanconi syndrome? A hereditary disorder of tubular functioWhat are some of the causes of tubular Fanconi syndrome, toxicity from compounds,

36


proteinuria? poisoning from heavy metals, ischemia due to obstruction of blood flow, decreased blood flow in shock or heart failure, toxicity from overload with proteins

What occurs when an increased load of small proteins (hemoglobin, myoglobin, free Ig light chain paraproteins) filtered through the glomerulus exceed the tubular uptake capacity?

Overload Proteinuria

What does detection of light chains depend on in overload proteinuria?

Electrophoresis and immunochemical testing

What color does hemoglobin or myoglobin confers to urine, yielding positive reactions in the dipstick test for blood?

Brown

What arises from injury, inflammation or malignancy of the lower urinary tract?

Postrenal Proteinuria

What does Postrenal proteinuria reveal under microscopic examination of urine?

Inflammatory and malignant cells and bacteria

What helps to distinguish whether a process is occurring in the kidneys or in the lower urinary tract?

Cast that arise in kidney tubules

What is an excellent screening tests for major glomerular injury (sensitive to albumin)?

Dipstick test

What is the extracellular fluid around the brain and spinal column?

Cerebrospinal fluid

Analysis of CSF is mainly performed to determine?

Increased permeability of blood-brain barrier (BBB) or increased intrathecal synthesis of immunoglobulin.

Which disease aids increased intrathecal synthesis of immunoglobulins?

Multiple sclerosis

Methods for measuring CSF protein includes? Pyrogallol red dye-binding assaysBenzethonium chloride turbidity assaysBiuret assays

What is analyzed in proteins in amniotic fluid? α-fetoprotein (AFP)What represents the fluid secreted by several salivary glands containing amylase and small peptides?

Proteins in saliva and oral fluid

Which immunoglobulin is tested in saliva for evaluation of possible immunological deficiency?

IgA

What is used in the diagnosis of gastrointestinal protein loss?

assay of feces for (α1-antitrypsin) AAT

What has been used as a measure of disease activity in inflammatory bowel disease?

Lactoferrin and calprotectin

Refers to proteins abnormally accumulated in the peritoneal (abdominal) and pleural (lung)

Proteins in Peritoneal and Pleural Fluid

37


cavitiesWhat are enzymes? Enzymes are organic catalysts than hasten a

chemical reaction without themselves being consumed or undergoing a chemical change.

How specific are they and where are the majority found?

They have a high degree of specificity for a certain substrate or class of substrates and themajority are intracellularly found.

What do the increases of serum enzymes usually indicate and why are they useful?

increased serum levels are due to tissue damage and necrosis, therefore they are used diagnostically to asses tissue damage.

What are Enzymes? are organic catalysts than hasten a chemical reaction without themselves being consumed or undergoing a chemical change

protein in nature with high degree of specificity for a certain substrate or class of substrates.

majority are intracellularly foundamounts found in serum are result of

normal cellular turnover; specialized tissue or widely distributed with tissue-specific isoenzymes or isoforms

increased serum levels are due to tissue damage and necrosis (markers of tissue damage)

thus, used diagnostically to assess tissue damage

What is necrosis? Necrosis is tissue damage due to elevating serum levels.

How many classes of Enzymes are there? There are 6 classes of Enzymes:I. Oxidoreductases

II. Aminotransferases/Aminotransaminases

III. HydrolasesIV. LyasesV. Isomerases

VI. LigasesWhat are the Classification of enzymes Oxidoreductase?

OxidaseCytochrome oxidaseDehydrogenaseLactate dehydrogenase (LDH)Malate dehydrogenase (MDH)Isocitrate dehydrogenase (ICD) Glucose-6-phosphate dehydrogenase (G-6-PD)

What are the Classification of enzymes Aminotransferase/Aminotransaminase?

Aspartate aminotransferase (AST / SGOT) Alanine aminotransferase (ALT / SGPT)

38


Creatine kinase (CK /CPK) Gamma glutamyl transferase (GGT) Ornithine carbamyl transferase (OCT)

What are the Classification of enzymes Hydrolase?

Esterase Acid phosphatase (ACP) Alkaline phosphatase (ALP) Cholinesterase (CHS) Lipase (LPS)Peptidases Typsin (PTS) Pepsin (PPS) Leucine aminopeptidase (LAP)Glucosidase Amylase (AMS) Amylo-1,6 glycosidase Galactosidase

What are the Classification of enzymes Lyases?

Aldolase (ALD) Carbonic anhydrase Glutamate decarboxylase Pyruvate decarboxylase Tryptophan decarboxylase

Two types of Isomerases: 1) Glucose phosphate isomerase2) Ribose phosphate isomerase

Another name for Ligases enzyme: Glutamine synthetaseThe roles of Oxidoreductases: a) Catalyze an oxidation-reduction

reactionb) Catalyze the addition or removal of

hydrogen from compoundsc) Require a coenzyme (NAD or NADP)

as a hydrogen acceptor or donor in order to function

The activities of Aminotransferases /Aminotransaminases:

a) Catalyze the transfer of amino or phosphate groups between compounds

b) Require coenzyme (pyridoxal-5’-phosphate [P-5’-P]) for the amino transfer reactions

The factors of Hydrolases: a) Catalyze the hydrolysis of ether and ester

b) They split molecules with the addition of water

c) Do not usually require coenzymes but often need activators

How do Lasses operate inside a cell? a) Catalyze the removal of groups without hydrolysis (loss of hyroxide ion)

39


b) Split molecules between carbon-to-carbon bonds without the addition of water

c) Resulting products usually contain carbon double bonds

How do Isomerases function? a) Catalyze the interconvertion of geometric or optical isomers

b) Catalyze the intramolecular conversions such as the oxidation of a functional group by an adjacent group within the same molecule

What are the tasks of Ligases? a) Catalyze the joining of two substrate molecules

b) Catalyze the union of two molecules accompanied by the breakdown of a phosphate bond in adenosine triphosphate (ATP)

The following are additional questions on slides 4-20How many classes of Enzymes are there? I. Oxidoreductases

II. Aminotransferases/Aminotransaminases

III. HydrolasesIV. LyasesV. Isomerases

VI. LigasesWhat are the classification of enzymes Oxidoreductase?

OxidaseCytochrome oxidase

Dehydrogenase Lactate dehydrogenase (LDH) Malate dehydrogenase (MDH) Isocitrate dehydrogenase (ICD)

Glucose-6-phosphate dehydrogenase (G-6-PD)

What are the classification of enzymes Aminotransferase/Aminotransaminases?

Aspartate aminotransferase (AST / SGOT)

Alanine aminotransferase (ALT / SGPT)

Creatine kinase (CK /CPK) Gamma glutamyl transferase (GGT) Ornithine carbamyl transferase (OCT)

What are the enzymes Hydrolases actions? Catalyze the hydrolysis of ether and esterthey split molecules with the addition of water.Do not usually require coenzymes but often need activators.

What are the enzymes Lyases actions? Catalyze the removal of groups without hydrolysis (loss of hyroxide ion).

40


Split molecules between carbon-to-carbon bonds without the addition of water.Resulting products usually contain carbon double bonds.

What are the enzymes Isomerases actions? Catalyze the interconvertion of geometric or optical isomers.Catalyze the intramolecular conversions such as the oxidation of a functional group by an adjacent group within the same molecule.

What are the enzymes Ligases Actions? Catalyze the joining of two substrate molecules.Catalyze the union of two molecules accompanied by the breakdown of a phosphate bond in adenosine triphosphate (ATP).

What is Phosphatase? is an enzyme that removes a phosphate group from its substrate by hydrolyzing phosphoric acid monoesters into a phosphate ion and a molecule with a free hydroxyl group.

What is Acid Phosphatase? Promotes the hydrolysis of orthophosphate esters.Found in almost all body tissues, but significantly in RBC’s and platelets.In adult men, 50% is found in the prostate gland.High levels are found in semen.

What are the requirement substrates to measure Total ACP Activity?

p-nitrophenylphosphate (PNPP) (preferred substrate for continuous monitoring)thymolphthalein monophosphate (TMP) (substrate of choice)

What is p-nitrophenylphosphate (PNPP)? Is a non-proteinaceous, non-specific substrate used to assay protein, alkaline and acid phosphatases.The PNPP phosphatase activity is measured using a continuous or single-point spectrophotometric assay based on the ability of phosphatases to catalyze the hydrolysis of PNPP to p-nitrophenol, a chromogenic product with absorbance at 405 nm (1).

What is thymolphthalein monophosphate (TMP)?

Obtained from 10 commercial suppliers were compared spectrophotometrically at 445 and 595 nm, liquid-chromatographically with monitoring at 254 nm, and enzymically by measurements of activity of prostatic acid phosphatase in human serum.

What is the measurement of Total ACP Activity?

All reactions are carried out at a pH lower than 6.0Falsely low values are produced by improper

41


anticoagulant use (heparin, fluoride, oxalate)Hemolysis produces false elevation

What is isoenzymes of Acid Phosphatase (ACP)?

Specific Prostatic Phosphatase Non-Specific Prostatic Phosphatase or Red

Cell PhosphataseWhat is Isoenzyme Test Include: inhibition of prostatic ACP by D-

tartrate, followed by the determination of the serum level of other ACP fractions

immunoassay with antibodies specific to certain isoenzymes

- electrophoresisWhat is the Clinical Significance of ACP? aids in detecting metastatic carcinoma

other types of cancer and bone diseases evidence of rape

What is Metastatic Carcinoma? Metastatic Carcinoma is the spreading of cancer, but with the Clinical Significant of ACP they can detect and prevent the cancer spreading.

What enzyme(s) have liver as the primary source and can be used to detect hepatic parenchymal disease?

Alanine aminotransferaseAspartate aminotransferase

What enzyme is a indicator in muscle disease ? Creatine kinaseWhat enzyme(s) associated with detecting pancreatic disease?

AmylaseLipase

Where would you find enzyme lipase ? PancreasWhat does ACP - Acid Phosphatase do? Promotes the hydrolysis of orthophosphate

esterWhere would you find ACP? In all body tissues, significantly in RBC’s and

plateletsIn adult men, where is 50% of ACP found? found in the prostate gland, high levels are

found in semenWhat condition the ACP activities were taken placed at?

pH lower than 6.0

What is D-

tartrate used in isoenzyme testing? inhibition of prostatic ACP , followed by the determination of the serum level of other ACP fractions

What are the clinical significance of ACP? aids in detecting metastatic carcinomaOther types of cancer and bone diseaseEvidence of rape

What is ALP alkaline phosphatase? Hydrolyzed phosphate estersWhat’s the optimal pH for ALP At 8.6 ÷ 10

What is ALP used for? diagnosis of hepatobiliary disease and bone disease associated with increased osteoblastic

42


Where will you find ALP? in liver, bone, kidney, intestines and placenta

What is the ALP isoenzyme that is most susceptible to heat inactivation (heat labile),?

Bone isoenzyme

What are bone disease patients will have elevated level of ALP?

- Osteomalacia- Rickets Paget’s disease

Why ALP level is high in children? It’s from the osteoblasts during bone growth

What are semiautomatic and automatic pipettes and dispensers used for?

These devices are programmed and used to simultaneously dispense aliquots into multiple wells.

Why are they piston driven? To allow the user to pipette with a few or as many tips as necessary.

What is centrifugation? The process of using centrifugal force to separate lighter from heavier parts of a solution, the correct term to describe the force required to separate two phases in a centrifuge is relative centrifugal force (RCF), also called relative centrifugal field.

Examples are horizontal head or swinging bucket, fix angle or angle head, ultracentrifuge and axial.

What is gravimetry? The process used to measure the mass of a substance using a balance to compare the mass of an unknown to that of a known mass. The comparison is called weighing and the absolute standards with which masses are compared are called weights.

What are the types of balances used? Double pan, single pan and the electronic balance.

The following are additional questions for slides 19:21-37What does Alkaline phosphatase (ALP) do and what is its optimum pH?

Hydrolyzes phosphate esters; optimum pH = 8.6-10

What are the activators for Alkaline phosphatase?

Activators include Mg2+, Co2+ and Mn2+

What are the inhibitors for Alkaline phosphatase?

Inhibitors include phosphate, borate, oxalate and cyanide ions

How is the Alkaline phosphatase used in clinical diagnosis?

Used in the diagnosis of hepatobiliary disease and bone disease associated with increased osteoblastic activity

Where Alkaline phosphatase is located? Found significantly in liver, bone, kidney, intestines and placenta

Does the bone isoenzymes of Alkaline phosphatase (ALP) tolerate to heat?

Bone isoenzyme; The most susceptible to heat inactivation (heat labile)

How does the bone isoenzymes Alkaline Elevated in persons with bone disease

43


phosphatase (ALP) influence in the person with bone disease?

a) Osteomalaciab) Ricketsc) Paget’s disease (osteitis deformans)

What is the outbreak of Alkaline Phosphatase (ALP) in children?

High in children; Alkaline phosphatase (ALP) from osteoblasts during bone growth

What is the B. Liver isoenzyme? It is one of the Five type isoenzymes of Alkaline phosphatase (ALP).

How does the B.Liver isoenzyme migrate on electrophoresis?

The most rapidly moving isoenzyme in electrophoresis

What is the outcome of B.Liver isoenzyme in person with hepatobiliary?

Greatly elevated in persons with hepatobiliary obstruction

What is the next type of isoenzymes Alkaline phosphatase (ALP)?

C. Intestinal isoenzyme

How does the C.Intestinal isoenzyme migrate on electrophoresis?

The slowest moving isoenzyme in electrophoresis

What is the D. Placental isoenzyme? D.Placental isoenzymes is belonging to Alkaline phosphatase (ALP) group.

What period does the D.Placental isoenzyme have a higher concentration?

Becomes elevated between the 16th and 20th weeks of pregnancy.

What is the optimum temperature for D.Placental isoenzyme?

The most heat stable (65°C)

What is the last type of Alkaline phosphatase (ALP)?

E. Regan isoenzyme

What is the carcinoplacental Alkaline phosphatase?

An abnormal Alkaline Phosphatase (ALP); A carcinoplacental Alkaline phosphatase (ALP) that has similar properties to the placental isoenzyme

What is the optimum temperature for carcinoplacental Alkaline phosphatase?

heat stable (65°C)

What is the method used to Measure of Alkaline phosphatase (ALP) activity?

Bessey-Lowry-Brock - methods

What is the substrate used in Bessey-Lowry-Brock-methods?

p-nitrophenylphosphate (PNPP)

What are the requirement cofactors in Bessey-Lowry-Brock-methods?

Required cofactors:Mg+2

Zn+2

What is the time window for Serum to be analyzed Measuring of Alkaline phosphatase (ALP) activity?

Serum (analyzed within 4 hours after collection) is preferred or heparinized plasma

What are the substances should be voided in Measurement of Alkaline phosphatase (ALP) activity?

EDTA, citrate and oxalate plasma must be avoided

Does the concentration of Alkaline phosphatase (ALP) in serum increase at refrigerated temperature?

Alkaline phosphatase (ALP) in serum stored at refrigerated temperature increases slowly (2%/day)

44


What should the frozen Alkaline phosphatase (ALP) be treated?

Frozen samples should be thawed 18-24 hours at room temperature

Does hemolysis increase results in Alkaline phosphatase activity?

Hemolysis falsely increases results

What is the most common method to differentiate Alkaline Phosphatase (ALP) isoenzymes?

Electrophoresis

How does Alkaline phosphatase (ALP) be placed on Electrophoresis?

Order of anodal migration:1) Liver ALP2) Bone ALP3) Placenta / Regan ALP4) Intestinal ALP

What substance will improve the separation of bone ALP and liver isoenzymes on Electrophoresis?

Pre-treatment with neuramidase for 15 minutes at 37°C improves separation of bone and liver isoenzymes

What residues must be dropped off bone ALP and liver ALP on Electrophoresis?

Removes the terminal sialic acid residues

Why should the terminal sialic acid residues be removed off Electrophoresis?

Sialic acid residues of bone ALP are more attached than liver ALP, thus reducing mobility

How does intestinal Alkaline phosphatase (ALP) be confirmed in serum?

Overnight incubation of serum with neuraminidase is used to confirm presence of intestinal Alkaline phosphatase (ALP)

Is intestinal Alkaline phosphatase (ALP) resistant to neuraminidase?

Intestinal Alkaline phosphatase (ALP) is neuraminidase-resistant

What is the second method to differentiate Alkaline phosphatase (ALP) isoenzymes?

Stability to denaturation by heat or chemical

Does Alkaline phosphatase (ALP) increase during the pregnancy?

Elevation during the 3rd trimester of pregnancy

What is the concentration of Alkaline phosphatase (ALP) in hepatitis and cirrhosis condition?

Elevation in hepatitis and cirrhosis

Does Alkaline phosphatase (ALP) increase in person with bone disease?

Elevation in bone diseases with osteoblastic activity

What will be happened to Alkaline phosphatase (ALP) in hyperthyroidism case?

Elevation in hyperthyroidism

How concentration of Alkaline phosphatase (ALP) in diabetes mellitus case?

Elevation in diabetes mellitus

What are the defintion for amiotransferases/(aminotransaminases)?

a) Aspartate Transferase (Transaminase) –AST

b) Serum Glutamate Oxaloacetate Transaminase –SGOT

How does the chemical reaction happened on Aspartate Transferase (AST)?

Transfers an amino group between as aspartate and keto acids

What is the first measurement of Aspartate The reaction with dinitrophenylhydrazine

45


Transferase (AST) activity? (DNPH) couples color reagent with keto acid product (Rietmann Frankel method)

What is the second measurement of Aspartate Transferase (AST) activity?

The reaction with diazonium salts couples the salt with the keto acid product and forms a color (Babson)

What is the third measurement of Aspartate Transferase (AST) activity?

The coupled enzyme assay (Karmen method) involves NADH, malate dehydrogenase and keto acid to form NAD. Malate dehydrogenase catalyzes the oxidation of oxaloacetate to malate in the indicator reaction.

What is the optimum temperature for Aspartate Transferase (AST)?

AST is stable in refrigerated temperature or frozen samples

What are the acceptable reagents in collection Serum or plasma Aspartate Transferase (AST)?

Serum or plasma collected with heparin, EDTA, citrate or oxalate is acceptable.

What substance is added to the Aspartate transferase (AST) specimen for stability?

Stability can be further enhanced by adding P-5’-P (pyridoxal-5’-phosphate) to the specimen

Where are Aspartate Transferase (AST) located?

AST is present in cardiac tissue, liver, skeletal muscles and RBC’s

Should Aspartate Transferase (AST) hemolysis specimens be avoided?

Hemolyzed specimens must be avoided

Does the alcohol affect Aspartate transferase (AST)?

Alcohol lowers AST values.

How many hours after onset of Myocardial infarction will increase the Aspartate transferase (AST)?

Elevated in:a) Myocardial infarctionb) Increases 6-8 hours after onset of pain

How fast Aspartate transferase (AST) will be elevated in Viral hepatitis?

Elevated in:a) Viral hepatitisb) 100-fold increase

Does Aspartate transferase (AST) increase its concentration in Skeletal muscle diseases?

Elevated in Skeletal muscle diseases likePolymyositisRhabdomyolysisDuchenne muscular dystrophy

Does Aspartate transferase (AST) affect Liver disease?

Elevated in:Liver hypoxia

What will be happened to the concentration of Aspartate transferase (AST) in strenuous physical activity?

Elevated in:Strenuous physical activity

What is the Aspartate transferase (AST) concentration in children?

AST tends to be higher in infants and children than adults

How does Aspartate transferase (AST) be classified in adult males and adult females?

Adult males exhibit higher AST activities than adult females

What are 5 clinical significance of ALP? 1) elevation during the 3rd trimester of pregnancy2) elevation in hepatitis and cirrhosis

46


3) elevation in bone diseases with osteoblastic activity4) elevation in hyperthyroidism5) elevation in diabetes mellitus

What does AST stand for? Aspartate TransferaseWhat does SGOT stand for? Serum Glutamate Oxaloacetate transminasewhat do AST and SGOT do? It transfers an amino group between an

aspartate and keto acids. L-aspartate + α-ketoglutarate AST oxaloacetate + L-glutamate

what are 3 major approaches involved in the measurement of AST activity?

1. The reaction with dinitrophenylhydrazine (DNPH) couples color reagent with keto acid product (Rietmann Frankel method)2. The reaction with diazonium salts couples the salt with the keto acid product and forms a color (Babson)3. The coupled enzyme assay (Karmen method) involves NADH, malate dehydrogenase and keto acid to form NAD. Malate dehydrogenase catalyzes the oxidation of oxaloacetate to malate in the indicator reaction.

AST is stable at what temperature? At refrigeration or frozen temperature.What specimen collection is needed for the measurement of AST?

Serum or plasma collected with heparin, EDTA, citrate or oxalate is acceptable.

What can you add to the specimen for AST so it can be more stable?

Stability can be further enhanced by adding P-5’-P (pyridoxal-5’-phosphate) to the specimen.

Where is AST present in the body? AST is present in cardiac tissue, liver, skeletal muscles and RBC’s.

What must be avoided in a specimen collected for AST measurement?

Hemolyzed specimens must be avoided.

What lowers the AST values? AlcoholWhat are the clinical significance of AST? AST is elevated in:

1) Myocardial infarction* Increases 6-8 hours after onset of pain2) Viral hepatitis* 100-fold increase3) Skeletal muscle disease* Polymyositis* Rhabdomyolysis* Duchenne muscular dystrophy

What is a micropipettes ? A hallow glass tube used for measuring small volumes of liquid in microliters and transfer of liquid from one source to anothere

47


Pipettes come in different sizes? Yes from single piece glass pipettes to more complex adjustable or electronic pipettes for various purposes with differing levels of accuracy and precision

What is the purpose of a pipette ? To draw up liquid for the purpose of measuring and transferring a specific volume of liquid

What is the mouthpiece of the pipette? The end of the pipette to which suction is applied to draw up liquid into the pipette

What is the calibration lines of a pipette? Marks on the stem of the pipette to show the point where liquid must be drawn to give a specific volume

What is Volume of the pipette ? Stated in millimeters example inscription may read 10ml in 1/10 ml and the pipette is graduated in 1/10 ml increments allowing it to be used to measure volumes up to 10 ml

What is the T.D of the pipette? “To deliver “pipettes designed to deliver a specific volume of fluid

What is the T. C of the pipette “To contain “ pipettes designed to contain a certain volume of fluid

Why must a distinction between T.D and T.C pipettes be made ?

Because there may be a difference between the volume a pipette will contain and a volume it will deliver

What is T.D calibrated with ? Distilled or deionized waterWhat is T.C glassware calibrated with ? MercuryWhat is the meniscus ? A crescent shaped structure appearing at the

surface of a liquid column it has the appearance of a contact lens

What is centrifugation ? The process of using centrifugal force to separate the lighter portion of a solution mixture or suspension from heavier portions.

What is gravimetric ? The process used to measure the mass of a substance

What is a Double-pan, single-pan and electronic balances

Types of balances

What are the Isoenzymes of CK? CK1 or CKBBCK2 or CKMBCK3 or CKMMCK- MtMacro- CK

Where is found CK1? brain and nerve tissuesWhere is found Ck2 in body? In cardiac muscles.At which level Ck2 is elevated and return to normal in MI?

levels rise within 4-6 hours, peak at 12-24 hours and return to normal within 48-72 hours

Where is found CK-Mt? Located between the inner and outer membranes of mitochondria, and it

48


constitutes up to 15% of total CK activityLocated on chromosome 15

How to measure CK by ion-exchange chromatography?

involves an anion exchange resin at a pH 8.0isoenzymes have different charges and can be selectively eluted

How to measure CK by immunoassay? involves antisera employed against certain subunits of each isoenzyme

How to measure CK by Tanzer-Gilvarg assay?

involves coupling with pyruvate kinase and lactate dehydrogenase to produce a change in absorbance measured spectrophotometrically.

How to measure CK by Oliver-Rosalki assay? involves coupling with hexokinase and G6PD, in which creatine is produced from creatine phosphate

Which are the Clinical Significance of CK? Increased in:-Myocardial infarction-Progressive Muscular Dystrophy (Duchenne type)-hypothyroidism-vigorous muscular activity (cerebrovascular accidents, stroke)-repeated intramuscular injections-drugs such as statins, fibrates, anti-retrovirals and angiotensin II receptor antagonists

Why should Avoid hemolysis in Measurement of CK activity?

Adenylate kinase, ATP and G6P/glucose-6-phosphate liberated from RBCs may affect the lag phase and may cause side reactions

The following are some additional questions on slides 38-54In what health condition/disease, the AST level found to be decreased ?

- Autoimmune hepatitis- Cholestasis- Cirrhosis- Alcohol liver disease

What does alanine aminotransferases? catalyzes the transfer of an amino group from alanine to α-ketoglutarate with the formation of glutamate and pyruvate

What is te required coenzyme for full catalytic activity of ALT?

Pyridoxal-5’-phosphate (P-5’-P)

How long can the sample being reserved samples stored at room temperature are stable for 3 days, 1 week if refrigerated

What is ALT:AST ratio (De Ritis ratio)used for?

determine the etiology of hepatocellular disease

What is ALT:AST ratio increased in? infectious (viral) hepatitisWhat is ALT:AST ratio decreased in? severe liver disease with necrosis (alcoholic

hepatitis, cirrhosis, hepatocellular carcinoma)

49


What enzyme Catalyzes the reversible phosphorylation of creatine by ATP

Creatine kinase

What is CREATINE PHOSPHOKINASE (CPK) or CREATINE KINASE (CK)

a dimer that is composed of two subunits: B-product and M-product

What are the isoenzymes of CK? - CK1 or CKBB- CK2 or CKMB- CK3 or CKMM

What appears in in brain damage or nerve tissues?

CK1 or CKBB

Where is CK2 located? Heart muscleWhat would be the optimal testing period for CKMB?

followingh MI, levels rise within 4-6 hours, peak at 12-24 hours and return to normal within 48-72 hours

What type of CK will be use as an indicative of iIschemic heart disease ?

- CK2 or CKMB

What is the indicative of myocardial damage? levels is >6%What enzyme is majorly found in striated and cardiac muscle and in normal serum?

CK3 or CKMM

What will CKMM find to be elevated in? hypothyroiidism and intramuscular injectionsWhere will CK-Mt find in? inner and outer membranes of mitochondria,

and it constitutes up to 15% of total CK activity

What can be used in Measurement of CK activity

- Electrophoresis (agarose, cellulose acetate)

- Ion-exchange chromatography- Immunoassay- Tanzer-Gilvarg assay (coulormetric

method)- Oliver-Rosalki assay

How to reserve CK for accurate result? - Refrigerated or frozen protected from light

- Addition of cysteineWhat substance is required as an activator for full enzyme activity? Mg

2+

In what condition would CK level increased? - Myocardial infarction- Progressive Muscular Dystrophy- Hypothyroidism- vigorous muscular activity

(cerebrovascular accidents, stroke)- repeated intramuscular injections- drugs such as statins, fibrates, anti-

retrovirals and angiotensin II receptor antagonists

50


What Catalyzes the conversion of lactate to pyruvate at pH 8.8-9.8 and pyruvate to lactate at 7.4-7.8, mediated by NAD?

Lactate dehydrogenase (LD/LDH)

What does LD stand for? Lactate DehydrogenaseWhat is the end product of LACTATE DEHYDROGENASE (LD/ LDH)?

pyruvate + NADH + H+

Which are the Isoenzymes of LDH?Which are the Isoenzymes of LDH? LD1 (HHHH; H4)LD2 (HHHM; H3M)LD3 (HHMM; H2M2)LD4 (HMMM; HM3)LD5 (MMMM; M4)

Where is found LD1 HHHH;H4 in body? predominantly in heart muscle and RBC’s

Where is found LD2 HHHM;H3M? localization is similar to LD1, but also found in the kidneys.

The Isoenzyme LD3 (HHMM;h2M2) is found where ?

lymphocytes, spleen and pancreas

The Isoenzyme LD3 (HHMM;h2M2) accounts for what percentage of total LD?

22%

The Isoenzyme LD4 (HMMM; HM3) is found where ?

Liver and Skeletal muscles

The Isoenzyme LD4 (HMMM; HM3) accounts for what percentage of total LD?

10%

LD5 (MMMM; M4) is similar to? LD4 in both localization and portion of totalBased on the detection of NADH the Wacker method uses?

the lactate-to-pyruvate reaction with the formation of NADH

Based on the detection of NADH the Wrobleuski-Ladue method employs what?

the reverse reaction of the Wacker method and measures the decrease in absorbance as NADH is consumed.

For measurement of Total LDH activity, why is serum the preferred specimen ?

-heparinized plasma also acceptable-oxalate has inhibitory effect on the enzyme

Measurement of Total LDH activity avoids what?

Hemolysis

For measurement of Total LDH activity, LD4

and LD5 can be stored at room temperature for how many days?

2-3 days

What is the normal LD1:LD2 ratio ? 0.45-0.74After MI, LD1 is > than LD2 (“flipped pattern”). Also observed in what?

Hemolyzed specimens

Immuno-inhibition for the measurement of LD1

activity involves what?an antibody that binds all other isoforms, leaving LD1 to be assayed by chemical reaction

Clinical Significance of LD is used for diagnosis of what?

Cardiac, hepatic, skeletal muscles and renal disease

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LD is elevated in association with what diseases ?

-Pernicious anemia-Viral hepatitis-Cirrhosis-Crush injuries-Renal disease

What are two clinical significances of LD1 and LD2 when elevated?

Hemolytic anemiaMyocardial infarction (MI)

What are three clinical significances of LD3 when elevated?

Pulmonary edemaPulmonary infarctionPancreatitis

What are two clinical significances of LD4 and LD5 when elevated?

Liver diseaseSkeletal muscle disease

The LD isoenzyme pattern in renal disease is very similar to a normal pattern except for the higher _____________?

Absolute values

Define Diagnostic Window. The interval of time after an episode of injury during which plasma concentrations of the marker are increased, thereby demonstrating the occurrence of injury

When does CK activity increase? 4-6 hours after the symptomsWhen does CK activity peak? 12-24 hoursWhen does CK activity return to normal? 48-72 hoursWhen does AST activity increase? 6-8 hours after the symptomsWhen does AST activity peak? 24-46 hoursWhen does AST activity return to normal? 4th of 5th dayWhen does LD activity increase? 8-12 hours after the symptomsWhen does LD activity peak? 48-60 hoursHow long does LD activity remain elevated? 10-14 daysWhat does AMI stand for? Acute Myocardial InfarctionWhat is a myoglobin? A heme protein that can bind oxygen

reversibly found in cardiac and striated muscles

How long does myoglobin take to increase after infarction?

1-3 hours

How long does myoglobin take to double its initial volume after infarction?

1-2 hours

When does myoglobin peak? 5-12 hoursWhen does myoglobin return to normal? 18-30 hoursWhat is myoglobin used in conjunction with to assess AMI?

Troponin and CK-MB

What is Troponin? A group of three proteins that function in muscle contraction by binding to the thin filaments of cardiac and skeletal muscles

What are the 3 types of Troponin? Troponin T (TnT)

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Troponin I (Tn I)Troponin C (TnC)

When does Cardiac troponin I (cTn I) increase after an AMI?

4-8 hours

How long does Cardiac troponin I (cTn I) remain elevated after an AMI?

3-5 days

When is a serial quantification of Cardiac troponin I (cTn I) necessary after an AMI?

3-6 hours6-9 hours12-24 hours interval

Where is Cardiac troponin T (cTnT) found in the body?

cTnT is found in regenerating and diseased skeletal muscle expressed in small quantities

What is GAMMA GLUTAMYL TRANSFERASE (GGT)?

GGT is an amino transferase that transfers a terminal gamma-glutamyl from one compound to an acceptor compound (transpeptidation)

What is an example of tissues containing significant amounts of GGT (descending order of activity per gram of tissue) ?

a. Kidneyb. Liverc. Pancreasd. Intestines

All of the above

E

How is GGT activity measured?a. Szasz Methodb. Nonhemolyzed serum

Both A and B

C

How is GGT activity stored? Stable for at least one month at 4°C and for one year at -20°C.

List the Clinical Significance of GGT:a. primarily used as a marker for the

diagnosis of liver related disorders (intrahepatic or post hepatic obstruction

b. detection of alcoholism and monitoring of alcohol intake

c. urine contains significant amount of GGT

d. elevated also in serum of patients receiving anticonvulsant drugs such as phenytoin and phenobarbital.

e. All of the above

E

Between male and female organism, who shows the higher GGT activities?

Male

GGT values are lowest in adolescents and rise steadily throughout life:

a. Trueb. False

A

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What does it usually mean when GGT and ALP enzymes are elevated and when GGT is normal but ALP is high?

If both enzymes are elevated, liver disorder is suspected;If GGT is normal, and ALP is high, suggestive of bone disease

What is Gamma Glutamyl Transferase (GGT)? Is an amino transferaseWhat does GGT do? Transfers a terminal gamma-glutamyl from one

compound to an acceptor compound (transpeptidation).

What contains a significant amount of GGT? TissuesName some tissues that contain GGT in descending order of activity per gram of tissue.

1) Kidney (proximal renal tubule)2) Liver3) Pancreas4) Intestines

What is used for the measurement of GGT? Szasz Method *substrate *gamma-Glutamyle-p-nitroanilide

What is the preferred specimen for GGT? Nonhemolyzed serum is the preferred specimen, but EDTA plasma has also been used.

What can decrease GGT”s activity by 10%-15%?

Heparin may produce turbidity; citrate, oxalate and fluoride.

In what temperature can GGT be stable in and for how long?

Stable for at least one month at 4°C and for one year at -20°C.

What diagnosis is GGT markers used for? Liver related disorders (intrahepatic or post hepatic obstruction).

What contains significant amounts of GGT? UrineWhat can the GGT assays detect? Alcoholism and monitoring of alcohol intake.In what other patients can GGT be elevated? In serum of patients receving acticonvulsant

drugs such as phenytoin and Phenobarbital.What are some clinical significance of GGT? 1) Acute pancreatitis

2) Diabetes mellitus (in relation to triglycerides increased)

Is GGT higher in males or females? In males (do to the contribution of GGT from the prostate gland)

GGT values are lowest in 1) _____? and 2) _____? steadily throughout life.

1) adolescents2) rise

What are GGT levels often examined with? ALPWhat is suspected if both enzymes (GGT and ALP) are elevated?

Liver disorder

If GGT is normal, and ALP is high then what is suspected?

Suggestive of bone disease

What is Amylase (AMS)? It is a digestive enzymeWhat is the optimum PH of AMS? 6.9-7.0What is and what does AMS do? a hydrolase that catalyzes the breakdown of

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starch and glycogen producing products consists of glucose, maltose and dextrins.

AMS is a metalloenzyme that requires what as a cofactor?

Ca2+

AMS is the _____? In size. SmallestWhat are the mist affective activators of AMS? Chloride and bromide

The only plasma enzyme physiologically found in urine is?

Amylase AMS

What are the 2 forms of AMS 1) P-type (pancreas) = ptyalin2) S-type (salivary glands) = amylopsin

What are 5 measurements of AMS activity? Amyloclastic method*measures amylase activity by following the decrease is substrate concentration *substrate = starch2. Saccharogenic method *measures the amount of reducing sugars produced by the hydrolysis of starch by the usual glucose methods3. Chronometric method *measures the time required for AMS to hydrolyzed completely all the starch present in a reaction mixture.4. Amylometric method *measures the amount of starch hydrolyzed in a fixed period of time using the intensity of the blue starch iodine color.5. Coupled-enzyme methods *measure change in absorbance of NAD. *AMS acts on maltopentose to produce maltose that is then reduced to form ATP. This reaction is the coupled with NAD.

When is the end point, in the chronometric method, reached?

When there is absence of any substrate capable of forming the blue starch iodine color.

What are some clinical significance of AMS? 1) Acute pancreatitis2) Salivary gland lesion3) Opiates medication

In acute pancreatitis, when do AMS levels rise?

5-8 hours after the onset of attack, peak at 12-72 hours, and then declines to normal in 3-5 days.

What are some salivary gland lesions? 1) Mumps2) Parotidis

Opiates medication causes pancreatic sphincters to do what?

Constrict

What is Macroamylasemia? An asymptomatic condition which results when the amylase molecule and

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immunoglobulins (IgG) combine to form a complex.

The complex is too large to be filtered across the?

Glomerulus

The lack of 1) ___ ___? leads to an 2) ____? serum amylase and 3) _____? Urine amylase.

1) Renal clearance2) Increased3) Decreased

What is evaluated using the ACCR (amylase: creatinine clearance ratio?)?

Macroamylasemia

What is ACCR? ACCR is the ratio of urinary clearance of amylase to urinary clearance of creatinine, expressed in %.

Macroamylasemia will show decreased urine amylase wit?

Low ACCR

In what is ACCR increased? In acute pancreatitisWhat is Lipase (LPS) A digestive enzyme secreted by the pancreas.LPS is filtered by what? The glomerulusLPS is totally reabsorbed by the? The renal tubules, thus is not physiologically

detected in urine.What does LPS do? It catalyzes the hydrolysis of ester bonds in

triglycerides to produce fatty acids and 2-acylglycerol.

What is used for the measurements of LPs activity?

*Generally, Titrimetric and Turbidimetric methods ~ Cherry Crandall ~ Tietz Templaton

Substrate = olive oil Cofactor = colipase

What is the preferred specimen for LPS assays?

Serum

What is the stability of LPS and at what temperatures?

1 week at room temperature 3weeks refrigerated 5 months frozen repeated freezing and thawing should

be avoidedWhat causes a reduction in lipase activity? 3. Lipemic specimensWhat increases lipase activity? 4. Opiates and morphineWhat are some clinical significance in LPS? 1. Acute pancreatitis

Lipase rises within 4-8 hours, peaks at about 24 hours, and decreases within 7-14 days.

2. Pancreatic duct obstruction3. Tumors of the pancreas4. Renal insufficiency

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From the graph below, the red line rises within 4-8 hours and peaks at about 24 hours, and decreases within 7-14 days, what is this digestive enzyme?

Lipase

The graph below indicates that the black line rise 5-8 hours after the onset of attack, peak at 12-72 hours, and then declines to normal in 3-5 days, what is this digestive enzyme?

Amylase

What do you call the enzyme synthesized by the liver that hydrolyzes the esters of choline?

CHOLINESTERASE (CHS)

What do you call an enzyme that is also known as “pseudo-cholinesterase”?

CHOLINESTERASE (CHS)

What does Acetylcholinesterase use as a substrate?

Acetlycholine

Where is Acetylcholinesterase found in the body?a. RBC, Spleenb. Gray matter, nerve endingsc. A and Bd. Stomach and Heart

C.

Acetylcholinesterase is the false cholinesterase or WBC cholinesterase. Evaluate this statement. True or False

False

Psuedocholinesterase uses what kind of substrate?

Butyryl esters (butyrylcholine)

Where is Psudeocholinesterase found in the body?a. serum, liverb. pancreas, heartc. gray matterd. A and B

D. A and B

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Pusedocholinesterase has other reference names. Which of these is false.a. serum cholinesterase (SChE)b. or acylcholine acylhydrolasec. butyrylcholinesterase, choline esterase

IId. acetylcholinesterase

D.

Manometric techniques measure liberated ____________ from the formation of acetic acid.

a. Carbon Dioxideb. Carbon Monoxidec. Phosphate Dioxind. Methane Gas

A

Electrometric techniques determine enzyme activity by measuring the ____ decrease resulting in the liberation of acetic acid.

a. Saltb. Basec. pHd. Aerosol

C

In the Ellman technique the substrate is a _______ ester that produces a ________, which reacts with a disulfide to form a colored compound.

a. Butryl, Butrylb. Thiol, thiolc. Glycol, glycold. Menthol, menthol

B

__________ is a commonly used substrate for measuring serum cholinesterase activity.

a. Psuedocholineb. Butrylcholinec. Acetylcholined. Propionylthiocholine

D

The Succinyldithiocholine method uses Serum or heparinized plasma only. The method is inhibited by__________.

a. Oxalateb. Fluoridec. Citrated. All the above

D

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PChe is stable for several weeks in refrigerated samples; __ years in frozen serum but only up to __ month(s) in frozen plasmaa. 3, 1b. 2, 2c. 4, 2c. 2, 6

A

When PChe is decreased it is clinically significant among other factors for

a. organic phosphorus insecticide poisoning

b. prolonged apnea after succinylcholine administration during surgery

c. hepatocellular disease, starvationd. All the above

D

_______ is an oxidoreductase that catalyzes the oxidation of glucose-6-phosphate to 6-phosphogluconate, which is an important step in the pentose-phosphate shunt in glucose metabolism.

G6PD

Name key organs where G6PD is found:a. Adrenal Glands, Thymusb. RBC’s, Lymph Nodesc. Stomach, Pancreasd. A and B

D

G6PD functions to maintain NADPH in a reduced form to protect hemoglobin from oxidation and prevent RBC hemolysis. True or False?

a. Trueb. False

A

The activity of G6PD is by the formation of NADPH which then is measured mass spectometrically.

True or Falsea. Trueb. False

B

G6PD is increased in Myocardial Infarction and Megaloblastic Anemia. True or False

a. Trueb. False

A

In Latinos G6PD is a common inherited sex-linked trait. True or False. a. True

A

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b.False

5’-NTP becomes elevated as a result of liver disease. True or False

a. Trueb. False

A

5’-NTP in _______ levels are more sensitive to liver cancer.

a. Serumb. Plasmac. A and Bd. blood

A

The lab wants to keep a sample stable for 5’-NTP. What is the longest temperature and time that it can be kept.

a. 4 days at 4 degrees centigradeb. 2 hours at room temperaturec. 4 months at minus 20 degrees

centigraded. None of the above

C

Lipoprotein-Associated Phospholipase A2 is a _______ risk marker.

a. Cardiovascularb. Endocrinec. Skeletal Muscular Systemd. None of the above

A

Which cells produce Lipoprotein-Associated Phospholipase A2

a. Monocytesb. Macrophagesc. Mast Cellsd. All of the Above

D

Lipoprotein-Associated Phospholipase A2 is used in detecting atherosclerotic lesions, especially in complex plaque prone to rupture. True or False

a. Trueb. False

A

Can increased levels of Lipoprotein-Associated Phospholipase A2 be associated with future coronary and cerebrovascular events?

a. Yesb. No

A

Lp-PLA2 can measured by the ELISA method and the Immunoturbidimetric method. True or False

A

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A. True B. FalseShould EDTA plasma be used as a recommended sample?

a. Yesb. No

A

Myeloperoxidase is released in circulation typically with inflammation.True or False

a. Trueb. False

A

Define Cancer. it is a term used for diseases in which abnormal cells divide without control and are able to invade other tissues. It can spread to other parts of the body through the blood and lymph systems.

DefineTumor Marker. Is a substance produced by a tumor or by the host in response to a tumor, which is used to differentiate the tumor from normal tissue, or to detect the presence of a tumor based on measurements in the blood or secretions

What is the use of Tumor Marker? It can be used for:--For diagnosis, prognosis and prediction For monitoring the effects of therapy -as targets for localization and therapy

What is Receiver operating characteristic curve (ROC) ?

It is a useful approach to evaluating multiple tests for the same analyte or multiple markers of the same type of cancer

Enzymes as Tumor Markers. Alkaline PhosphataseLactate DehydrogenaseNeuron-Specific EnolaseProstatic Acid PhosphataseProstate-Specific AntigenUrokinase-Plasminogen Activator SystemCathepsins

Hormones as Tumor Markers. Adrenocorticotropic hormone (ACTH)CalcitoninHuman Chorionic Gonadotropin (CG)

Oncofetal Antigens as tumor markers. α-Fetoprotein (AFP)Carcinoembryonic Antigen (CEA)

Cytokeratins as tumor markers. Tissue Polypeptide Antigen (TPA)Tissue Polypeptide Specific Antigen (TPS)CYFRA 21-1Squamous Cell Carcinoma Antigen (SCCA)

Carbohydrate Markers as tumor markers. -Mucins -CA 125-CA 15-3 -HE4-CA 27.29 -OVA1

Which are Bladder Cancer Markers? -Urinary Bladder Tumor Markers-Nuclear Matrix Protein (NMP22)

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-Bladder Tumor-Associated (BTA) Analytes

Proteins as tumor markers. -Soluble Mesothelin-Related Peptides (SMRPs)-Des-y-carboxy prothrombin (PIVKA-II)S-100 Proteins-Thyroglobulin and AntibodiesChromogranins

Receptors as tumor markers. -Estrogen and Progesterone Receptors-Epidermal Growth Factor Receptor

Genetic and Molecular Markers. Oncogenes:--RAS genes-HER2-BCR-ABLTumor-Suppressor Genes:--Retinoblastoma gene-adenomatous polyposis Coli (APC) Gene-BRCA1 and BRCA2

Which are the primary sources of Alkaline Phosphatase

liver, bone and placenta

What can be seen in primary and secondary liver cancer?

-Elevation of ALP with elevations in 5'-nucleotidase or γ-glutamyltransferase

When does Lactate dehydrogenase release take place?

-as a result of cell damage

Neuron-Specific Enolase (NSE) associated with which tumors?

tumors of neuroendocrine origin like small cell lung cancer (SCLC), neuroblastoma, pheochromocytoma, carcinoid, medullary carcinoma of the thyroid, melanoma and pancreatic endocrine tumor

What is Prostatic Acid Phosphatase (PAP)? It is considered to be less sensitive for the detection of early cancer but may prove useful when combined with other markers for improving prostate cancer detection or predicting recurrence after radical prostatectomy.

How is Prostate-Specific Antigen (PSA) produce?

produce exclusively by epithelial cells of the acini and ducts of the prostate gland and widely use to screen, detect and monitory treatment of prostate cancer.

Where is found Prostate-Specific Antigen (PSA)?

concentrations of PSA has been found to correlate with clinical stages of prostate cancer.

Which of the greatest clinical use of PSA? PSA involves monitoring of definitive treatment for prostate cancer which includes radical prostatectomy, radiation therapy and anti-androgen therapy.

Which methods are used to measure PSA? Sandwich immunoassays using labels such as enzymes, fluorescence or chemiluminescence

Which are the components mainly used in Urokinase-Plasminogen Activator System?

1. Urokinase plasminogen activator (uPA)2. uPA membrane-bound receptor (uPAR)

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3. uPA inhibitors, PAI-1 and PAI-2

Which is the main activity of Urokinase-Plasminogen Activator ?

the main activity is the conversion of plasminogen to active plasmin

What is the used of Urokinase-Plasminogen Activator System?

It has been used as a prognostic marker in breast cancer, colorectal cancer, ovarian, renal, hepatocellular, pancreatic, urinary, bladder, lung (adenocarcinoma) and cervical cancers as well as gliomas.

Which method is used to measure Urokinase-Plasminogen Activator System?

measured in the laboratory by the Enzyme-Linked Immunosorbent Assay (ELISA)

What is Cathepsin? Cathepsin are lysosome on protease enzymes: Cathepsins B, D and L

What happened if Cathepsin level increased? Cathepsin increased levels are demonstrated in breast, colorectal, gastric, lung, prostate carcinoma, gliomas, melanomas and osteoclastomas.

What methods are used to measure Cathepsin concentration?

Cathepsin concentrations are generally measured in tissue extracts by ELISA or directly in the tissues by immunohistochemistry

Which gland produced Adrenocorticotropic hormone?elevated

Adrenocorticotropic hormone produced by corticotropic cells of the anterior pituitary gland

What happened if Adrenocorticotropic hormone concentrations of plasma is elevated?

Elevated plasma concentration pancreatic, breast, gastric and colon cancer, and benign conditions such as chronic obstructive pulmonary disease, mental depression, obesity, hypertension, diabetes mellitus and stress.

Which are cells are responsible to produced Calcitonin?

It is produced by the C cells of the thyroid in response to increased serum calcium. It inhibits the release of calcium from bone and thus lowers the serum calcium concentration

What happened if Calcitonin concentration is elevated?

-Elevated concentration is usually associated with medullary carcinoma of the thyroid (MTC)-It is also elevated in patients with carcinoid tumors and cancers of the lung, breast, kidney and liver-It is also elevated in other nonmalignant conditions such as pulmonary disease, pancreatitis, hyperparathyroidism, pernicious anemia, Pagets' disease of bone and pregnancy

Which cells are produced Human chorionic gonadotropin (CG)?

Human chorionic gonadotropin (CG)is a glycoprotein secreted by the syncytiotrophoblastic cells of the healthy placenta

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What happened if Human chorionic gonadotropin (CG) elevated?

-Elevated concentrations are seen in pregnancy, trophoblast disease and germ cell tumors-Also elevated in biliary cancer, pancreatic cancer and other cancers including bladder, renal, prostate, liver, colorectal, non-small cell lung, breast and head and neck cancers

What is the use of GC and AFP together? GC together with α-fetoprotein (AFP) is useful in detecting nonseminomatous testicular tumor

What method is used to measure chorionic gonadotropin?

measured in the laboratory by a immunometric (sandwich) technique

What is Oncofetal antigens? Oncofetal antigensare proteins produced during fetal life

How to work Oncofetal antigens in fetal life? -present in high concentration in the sera of fetuses and decrease to low concentrations or disappear after birth-This proteins often reappear in cancer patients, indicating the reactivation of certain genes as a result of the malignant transformation of cells

Which are the Oncofetal antigens tumor markers? α-fetoprotein (AFP)Carcinoembryonic Antigen (CEA)

What happened if α-fetoprotein (AFP) is elevated? Elevated AFP and serum bilirubin concentrations (>2 mg/dL) are associated with a decreased survival time

α-fetoprotein (AFP) is determined by Which method?

Levels are determined by immunometric assay

What is the other name of Carcinoembryonic Antigen (CEA)?

"Gold" antigen

Which is the origin of Cytokeratins? Cytokeratins group of proteins that make up the cytoskeletal intermediate filaments of epithelial cells and cells of epithelial origin.

What happened if Tissue polypeptide antigen (TPA) is elevated?

Tissue polypeptide antigen (TPA)Elevated serum concentrations are related to the proliferative activity and turnover of cells, allowing it to be used as a proliferation marker.

What is the use of Tissue polypeptide antigen (TPA)?

In monitoring of metastatic disease, TPA is useful when comb (ined with CEA and CA 15-3 in breast cancer, with CEA and CA 19-9 in colon cancer, and with CA 125 in ovarian cancer.TPA may be helpful in the differentiation of cholangiocarcinoma (increased TPA) from hepatocellular carcinoma (in which TPA is not elevated)

What is tissue Polypeptide-Specific Antigen ? an antigenic site on cytokeratin 18 that is specifically recognized by the M3 monoclonal antibody

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CYTOKERATINS AS TUMOR MARKERS have been proposed as a specific marker for cell proliferation and is detectable in serum with the use of a _____ _____?

specific immunoassay

CYFRA 21-1 is elevated in all types of _____ _____, although it is most sensitive for non-small cell ____ _____.

lung cancer, lung cancer

Squamous Cell Carcinoma Antigen (SCCA) is a _______ referred to as tumor associated antigen 4.

glycoprotein

Squamous Cell Carcinoma Antigen (SCCA) is useful in detecting recurrence of ______ and in monitoring treatment and disease progression

cancer

CA 15-3 is detected by a _____ ______ _____ against a membrane-enriched extract of the human breast cancer metastatic to liver

murine monoclonal antibody

CA 15-3 elevated concentrations are found in ______ including pancreatic, lung, breast, ovarian, colorectal and liver cancers.

malignancies

CA 27.29 is used for detecting recurrent _____ _____ in patients with stage II or stage III disease and for monitoring response to therapy in patients with stage IV (metastatic) disease

breast cancer

CA 125 is a marker for monitoring what? epithelial ovarian cancerCA 125 is also used to detect ______ or _____ disease in patients who have undergone first-line therapy and would be considered for second-look procedures

residual or recurrent

CA 125 is also elevated in non-_____ carcinoma including pancreatic, lung, breast, colorectal, gastrointestinal tumors and useful in determining the prognosis of patients with endometrial carinoma

ovarian

CA 125 is determined by what? determined by immunoradiometric essayCA 19-9 (sialylated Lexa) a marker for gastrointestinal cancer and is used primarily in patients with _____ _____?

pancreatic carcinoma

CA 19-9 has been approved by the FDA for quantitative measurement in serum and as an aid in monitoring _____ _____ patient

pancreatic cancer

Monoclonal immunoglobulin has been used as a marker for multiple ______?

myeloma

Monoclonal paraproteins appear as sharp bands in the _____ region of serum electrophoretic

Globulin

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patterns.

Bence Jones proteins is what? a free monoclonal immunoglobulin light chain in the urine

What type of cancer is detected through cytoscopy or cytology of shed cells using tumor markers like NMP22, complement factor-H (CFH), fibronectin, telomerase, cytokeratins and survivin.

Bladder Cancer

Bladder cancer is measured what type of technique?

fluorescence in situ hybridization (FISH)

What type of proteins have functions that have been associated with regulating key reactions in the nucleus, such as DNA replication and RNA synthesis

Nuclear Matrix Protein (NMP22)

What detects the antigen human complement factor-H related protein, which is a variant of human complement factor-H

Bladder Tumor Associated analytes

Bladder Tumor Associated analytesfunctions in the alternative complement pathway by interacting with complement factor C3b to prevent what?

cell lysis

_______ is a cell surface glycoprotein express on mesothelial cells, and mesothelin fragments, found in the circulation of patients with mesothelial tumor.

Mesothelin

Mesothelioma is a rare cancer of the mesothelial surfaces of the pleural and peritoneal cavities or the pericardium that is linked to what?

asbestos exposure

What type of tumor markers are used as Calcium binding proteins for diagnostic histological marker of melanoma and melanoma metastases

S-100 Proteins

What type of protein is a family of proteins that are major components of the secretory granules of most neuroendocrine cells

Chromogranins

Chromogranins is useful in detecting _____ ______ like carcinoid tumor, pheochromocytoma and neuroblastoma

neuroendocrine tumors

Thyroglobulin and antibodiesproduced by the _____ gland as the precursor to ____ hormone

thyroid

Thyroglobulin and antibodies are used for monitoring patients with a diagnosis of differentiated ____ cancer after thyroid gland

Thyroid

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ablation.

What antibodies are used to monitor residual disease and or recurrence?

Antithyroglobulin

Thyroglobulin and antibodiesare measured by what?

Immunometric assays and RIA

Estrogen and progesterone receptors are members of the nuclear ____ hormone receptor family and are involved in hormone directed transcriptional activation

steroid

Estrogen and progesterone receptors are used in tumor markers as indicators of what type of cancer for hormonal therapy?

Breast cancer

Immunohistochemistry assay is used to measure steroid hormone receptors in what type of tumor tissue specimens?

breast

Epidermal Growth Factor Receptor (EGFR) is a prototype of what type of family receptors?

tyrosine kinase

What is used as a companion diagnostic to aid in identifying colorectal cancer patients?

Epidermal Growth Factor Receptor

Prostate Cancer Gene or Antigen3 (PCA3) a molecular test for what?

prostate cancer

Single-Nucleotide Polymorphismsgenes are associated with what?

human disease

Cell-Free Nucleic Acids are markers for what 2 types of cancer?

lung cancer and colorectal cancer

Creatinine is a diagnostic indicator of what function?

Kidney

What three organs is Creatinine synthesized in? Kidneys Liver Pancreas

True or False: Creatinine is present in all body fluids and secretions.

True

What is Creatinine a marker of? Glomerular Filtration Rate (GFR)What reaction is used in Creatinine? Jaffe reactionThe Jaffe reaction of Creatinine has a reaction with what?

Picrate ion

Creatinine analytical methods tend to lack what?

Specificity

Name two enzymes of Creatinine Creatininase creatinase

What are four analytical methodology factors that are used in Creatinine?

Enzymatic methods (Fig. 21-1)

Hydroxide Wavelength Temperature “Concentration”

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What type of state is Urea? • Azotemic stateUrea is an underestimation of what? • Glomerular filtration rateUrea is an outdated indicator of what function? • KidneyWhat is a chemical method of Urea? • Fearon reactionWhat is an enzymatic method of Urea based on?

• Preliminary hydrolysis of urea with urease

What three types of Urea enzymatic methods approached is used?

Spectrophotometric Dry Chemistry Systems• Specificity

What does a spectrophotometric approach consist of?

Berthelot method and enzymatic assay

What does the biochemistry and physiology of Uric Acid focus on?

Urate poolRenal handling of uric acid

A clinical significance of Uric Acid that involves, 20-plus inherited disorders of what metabolism?

Purine metabolism

Uric Acid is used as a measurement in management of what?

Gout

What does Uric Acid help determine on during Hypouricemia?

Link to underlying conditions Other causes

Name two analytical methodology of Uric Acid?

Phosphotungstic acid (PTA) Uricase

Which is more specific, PTA or Uricase? UricaseWhat does HPLC-based methods focus on during Uric Acid?

Ion-exchange or reversed-phase

What does reference values of Uric Acid indicate?

Gout Purine-containing and purine-free diets

What are Carbohydrates ? •Aldehyde or ketone derivatives of polyhydroxy (more than one -OH group) alcohols composed of carbon, hydrogen, and oxygen in the ratio of 1:2:1•they serve as structural components in RNA and DNA sugars•considered to be the main source of energy

What is a Monosaccharides ? Simple sugars that contain 4-8 carbons and only 1 aldehyde or ketone group.They are reducing sugars with the general formula of Cn(H2O) n that cannot be broken down to simpler substances by acid hydrolysis.

What is an example of a Monosaccharide? GlucoseWhat are Oligosaccharides ? Give an example Sometimes referred to as disaccharides which

are formed by the interaction of two monosaccharides with the loss of water molecule.Examples are: Maltose → two molecules of

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glucose;Lactose → glucose and galactose; Sucrose → glucose and fructose

Describe Polysaccharides? Carbohydrates formed by the interaction between many units of simple sugars.Examples include starch and glycogen

What is considered the primary energy source for the human body?

Glucose

What are the results of Glucose metabolism? •energy production by conversion to carbon dioxide and water•storage as glycogen in the liver or as triglycerides in adipose tissue•conversion to keto acids, amino acids or protein

What is Glycogenesis and Glycolysis? Glycogenesis is the conversion of glucose to glycogen and Glycolysis is the conversion of glucose or other hexoses into lactate or pyruvate.

What is Hypoglycemia and its symptoms? Hypoglycemia is a blood glucose concentration below the fasting value (<50 mg/dL or 2.75 mmol/L). Symptoms include trembling, sweating, nausea, rapid pulse, lightheadedness, hunger and upper abdominal discomfort

Describe neuroglycopenia and its characteristics

Neuroglycopenia is caused by the lack of glucose supply in the brain. It is characterized by headache, confusion, blurred vision, dizziness, seizures, loss of consciousness and even death.

How is a 72-hour test for Hypoglycemia conducted?

•Fasting hypoglycemia in adults can be monitored by the 72 hour fasting test•symptoms usually begin at plasma glucose concentrations below 55 mg/dL, and impairment of cerebral function begins when glucose is less than 50 mg/dL•the 72 hour fast should be conducted in the hospital•the patient should be allowed a liberal intake of calorie free and caffeine free fluids

How to determine a true Hypoglycemia? True hypoglycemia will show an abnormally low value within 12 hours of beginning a fast

How does Ethanol produces hypoglycemia? Ethanol produces hypoglycemia by inhibiting gluconeogenesis, and this inhibition is aggravated by malnutrition (low glycogen stores) in individuals with chronic alcoholism.

One way to determine an insulin producing pancreatic islet cell tumor is?

The demonstration of a low plasma glucose concentration in the presence of a high plasma

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insulin value

Name possible causes of Postprandial Hypoglycemia

•drugs•antibodies to insulin or insulin receptor•inborn errors (fructose-1,6- diphosphatase deficiency)•reactive hypoglycemia (functional hypoglycemia)

What are some criteria of Whipple's triad overflow blood glucose concentration?

•symptoms known or likely to be caused by hypoglycemia•low glucose when symptoms occur•relief of symptoms when glucose is increased the normal

What are the Pathophysiological mechanisms that contribute to hypoglycemia in patients with diabetes?

Defective glucose counter regulation (ability to increase glucose, counter to the effect of insulin)unaware of hypoglycemia

Where does the Lactic acid predominantly derived from?

white skeletal muscle, brain, skin, renal medulla and erythrocytes, and are metabolized in the liver and kidneys

What is the Cori cycle ? The Cori cycle is the conversion of glucose to lactate in the periphery and reconversion of lactate to glucose in the liver

How can the measurement of pyruvate be useful?

Measurement of pyruvate is useful in the evaluation of patients with inborn error of metabolism who have increase serum lactate concentration.Pyruvate is also measured in clinical studies evaluating reperfusion after myocardial ischemia

What can a lactate-to-pyruvate ratio indicate? A lactate-to-pyruvate ratio of less than 25 suggests a defect in gluconeogenesis, whereas an increase ratio (<35) indicates reduced intercellular conditions found in hypoxia

Describe Lactic acidosis Lactic acidosis is a lactate concentrations exceeding 5mmol/L with pH less than 7.25.Occurs in 2 clinical settings:•Type A (hypoxic) associated with decreased tissue oxygenation•Type B (metabolic) associated with diabetes mellitus, neoplasia, liver disease, drugs and inborn error of metabolism

When testing for carbohydrate metabolism, a deficiency or absence of an enzyme that participates in carbohydrate metabolism may

Accumulation of monosaccharides, which are measured in the urine

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result in what inborn error?Name the 2 sugars of clinical interest that are found in the urine?

Glucose and Galactose

Glucose can be checked by what? Glucose oxidase stripsWhy should urine from infants and children be tested by both the glucose oxidase and copper reduction test?

To identify individuals with inborn error of metabolism

Name the 5 techniques used to separate and identify sugars?

fermentation optical rotation osazone formation with

phenylhydrazine specific chemical test paper or thin layer chromatography

Where in the body is glycogen predominantly stored?

Liver and Skeletal muscles

What is the cause of Glycogen Storage Disease?

Deficiency of a specific enzyme in glycogenmetabolism

Lipids are a class of compounds that are soluble in but insolublein ?

Organic solvents; water

Which organic compound can serve as hormones, energy source, assist with ingestion and act as structural components in cell membranes?

Lipids

Name the 6 forms of lipids: Cholesterol Fatty acids Acylglycerols (Glycerol esters) Sphingolipids Prostaglandins Terpenes

What form of lipid is an unsaturated steroid alcohol whose structure is based on the CPPP nucleus?

Cholesterol

CPPP stands for? CyclopentanoperhydrophenanthreneWhere does absorption of cholesterol mostly occur?

In the middle jejunum and the terminal ileum of thesmall intestines

Absorption of cholesterol is mediated by what enterocyte protein?

NPC1L1 (Niemann-Pick C1-like1).

In the intestinal mucosal cells, cholesterol is packaged with triglycerides, phospholipids and a large protein, called apolipoprotein (apo) B-48, into large lipoprotein particles called?

Chylomicrons

What is the function of the chylomicrons? Chylomicrons are secreted into the lymph and enter the circulation, where they deliver the absorbed dietary lipid to the liver and peripheral tissues

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Cholesterol is synthesized by all cells in the body, but particularly by what organs?

Liver and intestines

What are the three stages to cholesterol biosynthesis?

involvement of acetyl-coenzyme A (CoA)

involvement of the microsomal enzyme HMG-CoA ( hydroxy-methyl-glutaryl CoA) reductase

a series of oxidation-decarboxylation reactions in the endoplasmic reticulum

Cholesterol is esterified to a fatty acid to form a cholesteryl ester by what two enzymes?

acylcholesterol acyltransferase (ACAT) lecithin cholestrol acyltransferase

(LCAT)What happens to cholesterol when it is esterfied?

It loses its free hydroxyl group and becomes more hydrophobic

Specialized endocrine cells can use cholesterol for the synthesis of what?

Steroid hormones

How is most cholesterol catabolized? Most cholesterol is converted in the liver into bile acids

What is referred by the term, essential fatty acids?

The term "essential fatty acid" refers to fatty acids required for biological processes but does not include the fats that only act as fuel.

Name the only 2 fatty acids known to be essential to humans:

alpha-linolenic acid (an omega-3 fatty acid)

linoleic acid (an omega-6 fatty acid)What is the term that is used for other fatty acids that are sometimes essential under some developmental or disease conditions?

Conditionally essential

What 2 fatty acids are conditionally essential? docosahexaenoic acid gamma-linolenic acid

Where does the fatty acid get metabolized and produced energy by a series of reactions known as β-oxidation?

Mitochondria

When fatty acid is metabolized it produces energy by a series of reactions, this reaction is called what?

β-oxidation

What is Krebs cycle? A common pathway for the final oxidation of nearly all metabolic fuels (carbohydrate, fat or protein) and ultimately results in the production of ATP (adenosine triphosphate)

What is the main energy storage molecule in the body?

ATP (adenosine triphosphate),

True or FalseKetone formation develops from excessive production of acetyl-CoA (ketosis)

True

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True or FalseIn ketone formation, as the body attempts to obtain necessary energy from stored fat, in the absence of an adequate supply of carbohydrate metabolites as in cases of prolonged starvation or when carbohydrate metabolism is impaired (as in diabetes mellitus)

True

What are the forms of ketone bodies? acetoacetic acidΒ-hydroxybutyric acidacetone

What do you call a three-carbon alcohol that contains a hydroxyl group on each of its carbon and is classified by the number fatty acyl groups present?

ACYLGLYCEROLS (GLYCEROL ESTERS)

What is acylglycerol? A three-carbon alcohol that contains a hydroxyl group on each of its carbon and is classified by the number fatty acyl groups present: one fatty acid - monoacylglycerols (monoglycerides); two fatty acids - diacylglycerols (diglycerides); and three fatty acids - triacylglycerols (triglycerides)?

How many carbons does ACYLGLYCEROLS contain?

3

What do you call an ACYLGLYCEROL that has one fatty acid?

monoacylglycerols

What do you call an ACYLGLYCEROL that has two fatty acids?

diacylglycerols (diglycerides);

What do you call an ACYLGLYCEROL that has three fatty acids?

triacylglycerols (triglycerides)

What type of lipid that constitutes 95% of tissue storage fat and are the predominant form of glyceryl esters found in plasma?

Triglycerides

Triglycerides from plant sources (corn, sunflower, safflower) are liquid oils at room temperature tend to be enriched with what kind of fatty acid?

Unsaturated fatty acids

Triglycerides from animals (ruminants) that are solid at room temperature and tend to have this kind of fatty acid, what is this fatty acid called?

saturated fatty acids

Where does dietary triglyceride get digested? Dietary triglycerides are digested in the duodenum

After being digested in the duodenum, dietary triglycerides get absorbed where?

Absorbed in the proximal ileum.

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Dietary triglycerides through the action of pancreatic and intestinal lipases and in the presence of bile acids, they are first hydrolyzed to what?

To glycerol, monoglycerides, and fatty acids.

Through the action of and intestinal lipases and in the presence of bile acids, they are first hydrolyzed to ____(1)___, ___(2)___,and ___(3)_. After absorption, they are reassembled as triglycerides in the intestinal epithelial cells and then are package with cholesterol and apoB-48 to form ___(4)_____

1) glycerol2) monoglycerides3) Fatty acids4)chylomicrons

What do you call the lipoprotein that carries triglycerides to the liver and peripheral cells?

chylomicrons

______________are the main metabolic fuel carried by chylomicrons to the liver and peripheral cells.

Triglycerides

A ________ is a dihydric 18-carbon alcohol, derived from the amino alcohol sphingosine.

Sphingolipids

What form of lipids that have a more general role in cellular interactions and serves as a source of blood group and tumor antigens?

Sphingolipids

What are prostaglandins? Any of the group of compounds derived from unsaturated 20-carbon fatty acids (primarily arachidonic acid) via the cyclo-oxygenase pathway

What group of physiologically active lipid compounds, which have diverse hormone-like effects in animals and found in almost every tissue in humans and other animals?

prostaglandins

Prostaglandins are derived enzymatically from?

fatty acids

True or FalseThe structural differences between prostaglandins account for their different biological activities.

True

True or FalseA given prostaglandin may have different and even opposite effects in different tissues.

True

The ability of the same prostaglandin to stimulate a reaction in one tissue and inhibit the same reaction in another tissue is determined by?

The type of receptor which the prostaglandins bind.

With their target cells present in the immediate vicinity of the site of their secretion, Prostaglandin acts as what?

autocrine or paracrine factors

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How does Prostaglandins differ from endocrine hormones?

Prostaglandins differ from endocrine hormones in that they are not produced at a specific site but in many places throughout the human body.

What are the two derivatives of prostaglandin? prostacyclins and thromboxanesWhat derivative of prostaglandin that is powerful locally acting vasodilators and inhibits the aggregation of blood platelets?

Prostacyclins

Through their role of this prostaglandin derivative in vasodilation, they are also involved in inflammation. What is this derivative called?

Prostacyclins

Where does Prostacyclins get synthesized? They are synthesized in the walls of blood vessels and serve the physiological function of preventing needless clot formation, as well as regulating the contraction of smooth muscle tissue

What derivative of prostaglandins produced by platelet cells?

Thromboxanes

What Prostaglandin derivatives are vasoconstrictors and facilitate platelet aggregation?

Thromboxanes

What is this polymer consisting of the five-carbon isoprene unit and include vitamins A, E, and K and the dolichols, which play important roles in protein glycation?

Terpenes

_____(1)____ and _____(2)______ are the primary constituents of the essential oils of many types of plants and flowers. Essential oils are used widely as fragrances in perfumery, and in medicine and alternative medicines such as aromatherapy.

1)Terpenes2)terpenoids

What are these macromolecular complexes that serve as transport vehicle of insoluble lipids in the plasma?

Lipoprotein

What is the role of the lipoprotein in insoluble lipids in the plasma?

Serves as transport vehicle of insoluble lipids in the plasma.

What is the appearance of a lipoprotein? They are typically spherical particles with nonpolar neutral lipids (triglycerides and cholesterol esters) in their core and more amphipathic lipids (phospholipids and free cholesterol) at their surface.

What do you call this major protein component that is found on the surface of a lipoprotein?

Apolipoproteins

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What are the categories of lipoprotein? 1)Chylomicrons2)Very Low Density Lipoprotein (VLDL) or pre-β-Lipoprotein3)Intermediate Density Lipoprotein (IDL)4)Low Density Lipoprotein (LDL) or β-Lipoprotein5)High Density Lipoprotein (HDL) or α-Lipoprotein6)Lipoprotein (a) [Lp(a)]

True of False.The larger lipoproteins contain more core lipids, triglycerides and cholesterol esters and are lighter in density and contain a small percentage of protein.

True

True or FalseIn the fasting state, most plasma triglycerides are present in VLDL, but 10 to 12 hours after a meal, most triglycerides are transported on chylomicrons.

False (2 – 6 hours after a meal)

How much total of plasma cholesterol is carried by LDL?

carries about 70% of total plasma cholesterol

What Lipoprotein contains about 20% to 30% of plasma cholesterol?

HDL

True or FalseLower density corresponds to larger particle and lower protein-to-lipid ratio, while higher density are smaller and have higher proportions of protein

True

In lipoprotein, density is inversely proportional to the protein content and directly proportional to the lipid content and size of the molecule

False (directly proportional to the protein content and inversely proportional to the lipid content and size of the molecule)

Complete the chart:_(1)_ density = _(2)_ protein content↑ density = _(3)_ lipid content_(4)_ density = ↓ size of the molecule (smaller)

1) ↑2) ↑3) ↓4) ↑

Are the Apolipoproteins; major components of lipoproteins?

True or FalseA. TrueB. False

A

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Apolilipoproteins function to:A. modulate the activity of enzymes that

act on lipoproteinsB. maintain the structural integrity of the

lipoprotein complexC. facilitate the uptake of lipoprotein by

acting as ligands for a specific cell-surface receptors

D. All of the above

D

Apo A-1 is not the major protein in HDL. True or False

A. TrueB. False

B

Chylomicrons have what type of Apolipoprotein? A. Apo A-1 B. Apo B-100 C. Apo B-48 D. none of the above

C

Lipoproteins have several pathways. Identify them.

A. Exogenous Pathway and the Endogenous Pathway

B. Intercellular-Cholesterol Transport Pathway and the Reverse-Cholesterol Transport Pathway

C. A and BD. None of the Above

C

The transport of dietary lipids from the intestines to the liver and peripheral cells mediated by:

A. ChylomicronsB. RBC’sC. WBC’sD. Adipose Tissues

A

The endogenous pathway’s is transfer _______ derived________ .

A. Hepatically, LipidsB. Splenically, TriglyceridesC. Kidney, MonoglyceridesD. Stomach, Diglycerides

A

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The endogenous pathway has which mediator lipoprotein? A. Apo A-1 B. Apo B-100 C. Apo B-48

D. all of the above

B

Cells use the cholesterol for several roles. Identify the appropriate role.

A. Membrane BiogenesisB. Vesicular TransportC. OsmosisD. Exocytosis

A

Do cells have a surface receptor for cholesterol?

A. YesB. No

A

The Reverse Cholesterol Transport Pathway’s function is to remove excess cellular cholesterol from peripheral cells and return it to the liver for excretion. What is the mediator?

A. HDLB. VLDLC. LDLD. chylomicrons

A

Are Coronary Heart Disease, Type V Hyperlipoproteinemia, and Familial hypercholesterolemia clinically significant?A. YesB. No

A

The Lab has been asked to conduct a total lipid panel. What tests must be run?

A. Total CholesterolB. TriglycerideC. HDL-CD. LDL-CE. All of the above

E

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