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Basic Examination of BloodBlood is an important specimen in the clinical laboratory. The importance of blood in medical branches:

• Microbiology: to detect systemic infection of bacteria through immunoassay in blood culture.

• Liver and other organ: specific enzyme in each site of organ can be warning for damaged tissue/organ (not normally found in blood/low in blood)

• Hematology: abnormal cell count and concentration, morphology can indicate blood disease and disorder (anemia, leukemia, thrombopenia, malignancy)

• Indication of certain disease: disorder, deficiency, or toxicity. e.g.: Blood glucose increase = diabetes mellitus Cholesterol, LDL, triglyceride increase = obese, cardiac failure Blood protein decrease = malnutrition, kwashiorkor

- Pharmacology: response of drug - Radiology: response of radiation

Advantages: Easy to collect, carries important info about patient’s condition, many tests can be carried out

COMPLETE BLOOD COUNT (CBC) Provides important information about the kinds and numbers of RBC, WBC, platelet Parts of routine physical examination Help to: Evaluate symptoms (weakness, bruising, fever, weight loss)

Diagnose condition (anemia, infection) Diagnose diseases of the blood (leukemia) Monitor response to some types of

drug/radiation treatmentResults: Normal value can vary from lab to lab. Normal value for CBC tests vary, depending on age, sex, elevation above sea level, and type of sample.

1. HemoglobinThe main component of RBC, gives RBC it’s red color. Conjugated protein; contain 2 pairs of polypeptide (globin) and 4 prosthetic heme group.

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Main function: transport O2 from lungs (conjugated protein: vehicle for transport of O2 & CO2).Derivatives of hemoglobin: hemiglobin (methemoglobin), sulfhemoglobin, carboxyhemoglobin (most)

Examination aim: measure the amount of hemoglobin in blood

Clinical implication: E.g. Hb low → anemia Determining concentration of Hb with

Hemiglobincyanide (HbCN) method *hazard to environment

Principle: Blood is diluted in a solution of potassium ferricyanide (K3Fe(CN)6) then measured by spectrophotometer at 540 nm and compared with that of a standard Hi CN solution.

Errors in hemoglobinometry : error inherent in the sample, method, equipment, operator’s error

2. HematocritExplanation: ratio of volume of erythrocytes to whole blood. Expressed in % (conventional) or decimal fraction L/L. (SI units)

Venous puncture shows same result as skin puncture → both greater than body hematocrite. It reflects concentration of red cells not total cell mass Clinical implication: green/orange plasma:

hemoglobinemia(rising number of Hb)

low Ht = anemia, high Ht= polycythemia

Method & units: mix thoroughly before taking sample Direct: centrifugation with macro/micromethod Indirect: automated instrument → MCV x RBC count

Sources of error: centrifugation, sample, etc.

3. Blood cell counting Expressed as concentration – cells per unit volume

of blood (mm3/μL) CBC can be performed:• Manually :

– Hemocytometer– Calculate from other CBC results (RBC

indices)• Semiautomatically• Automatically (hematologic analyzer)

a) WBC count

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To calculate number of components of WBC; help to identify infection (neutrophilia, lymphocytosis) and allergic or toxic reaction to certain medication (eosinophilia)

Expressed as concentration – cells per unit volume of blood (mm3)

Components: PMN, band neutrophil, lymphocyte, monocyte, eosinophil, basophil

Principle: anticoagulant EDTA + diluting fluid to lyse.

Clinical aspect: high WBC=leukocytosis, low WBC = leucopenia.

b) WBC differential counts Measure the percentage of each type of white

blood cells Components: PMN, band neutrophil (include

immature neutrophil), lymphocyte, monocyte, eosinophil, basophil

Expressed as a percentage of each type Increase/decrease number of each type help to

identify : Infection (neutrophilia, lymphocytosis); Allergic or toxic reaction to certain medication (eosinophil/basophil); Malignancy (leukemia)

c) RBC cell count Count number of RBC in every unit of blood

volume

Expressed as concentration – cells per unit volume of blood (mm3).

Clinical implication: < normal → anemia, higher → polycythaemia

Methods & units: manual → hemocytometer chamber, can use semiautomated method

d) RBC indices Determine the size of RBC, Hb content and

concentration in the RBC Useful for morphologic characterization of anemia Maybe calculated from: Red cell count, Hb

concentration, or Hematocrit 3 RBC indices :

MCV (Mean Corpuscular Volume)• Shows the size of RBCs, average volume of RBC• Classified as: normocyte, microcyte, macrocyte• Calculated from the Ht and RBC count• Expressed in femtoliter (fL) or cubic

micrometers• Diurnal variation: highest in the morning, lowest

in the evening

MCH (Mean Corpuscular Hemoglobin)

MCV = Ht ×1000 / RBC (in million per μL)

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• Is the content (weight) of Hb of the average RBCs.

• Calculated from the Hb concentration and RBC count.

• Expressed in picograms (μμg) or pg• Clinical implication: MCH low=Hypochromic RBC,

MCH high= Hyperchromic RBC (reticulocyte)

MCHC (Mean Corpuscular Hemoglobin Concentration)• Is the average concentration of Hb in a given

volume of packed red cells• Calculated from Hb concentration and the

hematocrit. • Expressed in g/dL• Clinical implication: MCHC low: Hypochromic

RBC

e) Platelet countPlatelets normally are 2-4 μm in diameter, 5-7 fL in volumeFunction: Homeostasis, maintain vascular integrity, blood clotting, more difficult to count (small, tendency to adhere to glass and one another), MPV (Mean

Platelet Volume) can also be counted, women have greater number of platelet than men Is expressed as concentration – cells per unit

volume of blood (mm3) Clinical implication: platelet increase:

thrombocytosis; decrease: thrombocytopenia

** Reticulocyte count • Characteristic: immature non-nucleated red cells, contain RNA, continue to synthesze Hb after loss of nucleus • Clinical implication: increase reticulocyte=increase production of RBC

4. Blood film examination Principle: blood spread in glass slide/covering Aim: to evaluate the numbers, size, and shape of

RBC, WBC and platelet in a stained smear of peripheral blood

Red Cell Distribution W idth (RDW) • Reports whether all the red cells are about the

same width, size, and shape.• This helps further classify the types of anemia.

5. Erythrocyte Sedimentation Rate (ESR)

MCH = Hb (g/L) /RBC (in million per μL)

MCHC = Hb (g/dL) /Hct

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Useful, non-specific marker of inflammation Measures:

- The settling of erythrocytes in diluted human plasma over a specified time period (1 hour)

- Distance from the bottom of the surface meniscus to the top of erythrocyte sedimentation in a vertical column containing diluted whole blood.

Normal values :Adult men 0-15 mm/hAdult women 0-20 mm/h

Methods:- Manual :

o Westergren Methodo Wintrobe & Landsberg Method

- Automatic

Factors affecting:- RBC size & shape

Rouleaux → increase ESRRate: microcyte → macrocyteSpherocyte → decrease ESRSickle cell → decrease ESR

- Plasma fibrinogen & globulin levelsPresence of fibrinogen → increase ESR

Fibrinogen decrease (-) charge (zeta potential) of erythrocytes that keep them apart. If zeta potential decrease, rouleaux will be formed. Excess immunoglobulin → increase ESR.

- Mechanical- Technical

Conditions associated with an elevated ESRRheumatoid arthritis, Multiple Myeloma, Cryoglobulinemia, Temporal arteritis, inflammatory diseases, Pregnancy, Anemia, Malignant neoplasms, Paraproteinemias, Macroglobulinemia, Hyperfibrinogenemia, Chronic infections, Collagen disease, Polymyalgia rheumatica

OTHER EXAMINATIONS USING BLOOD SPECIMEN

Liver function tests are one of the blood tests that are most commonly performed to assess the function of the liver or injury caused to the liver. Liver

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damage is detected initially by performing a simple blood test that determines the level of various liver enzymes present in the blood. The most widely used liver enzymes that are sensitive to abnormalities in liver and are most commonly measured are the aminotransferases. The two aminotransferases that are checked are the alanine aminotransferase (ALT or SGPT) serum glutamic pyruvic transaminase and aspartate aminotransferase (AST or SGOT) serum glutamic oxaloacetic transaminase.

SGOT and SGPT are highly sensitive markers of liver damage due to various diseases or injury. However, the fact is that higher than normal levels should not be automatically considered as indicative of liver damage. They may or may not imply liver disease. For instance, these enzymes are also elevated in cases of muscle damage.

SGOT Normally present in a number of tissues such as

heart, liver, muscle, brain and kidney. It is released into the blood stream whenever any of these tissues gets damaged.

Blood AST level is increased in conditions of muscle injury and heart attacks. Hence, it is not highly

specific liver tissue damage indicator as it can be elevated in conditions other than liver damage.

The normal levels of SGOT is in between 5 and 40 units per liter of serum.

SGPT Normally present in large concentrations in the

liver. Hence, due to liver damage its level in the blood rises, thereby, serving as a specific indicator for liver injury.

The normal levels of SGPT in between 7 and 56 units per liter of serum.

However the normal ranges of SGPT and SGOT differ depending on the protocols and technique used to measure in laboratory.