English Language and Foundation Studies Centre

EPBIOL259/EPCHEM314 Life Sciences

Laboratory Session

Part A: Using a Microscope

Part B: BMR and Cardiovascular Physiology

Student Name:

Student Number:

Course: (please circle)

EPBIOL259 EPCHEM314

Marking:

………………./10 Demonstrator Signature:

Life Sciences Laboratory Session

Marks from this assessment will contribute towards your final grade for this course. Please complete all working and answers on this sheet. This assessment must be submitted before leaving the laboratory.

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Conducting an experiment in a laboratory involving glassware, equipment and other people requires skill and patience. The most important thing is for you to be fully prepared. Always read carefully ahead before beginning, try to visualise the steps, write down any questions and clear them with the instructor before you begin. Safety Issues and Requirements:

1. All students must wear covered footwear; entry to the laboratory will not be permitted otherwise. Long hair must be tied back, no bare midriffs.

2. Eating, drinking or smoking is not permitted in the laboratory. 3. Any sharps, broken glass must be placed in the special containers provided. 4. Only supervised work is permitted in the lab. 5. Any spills or accidents must be reported immediately. 6. Consult the MSDS for unfamiliar chemicals. 7. Wash your hands immediately after leaving the laboratory. 8. If in doubt always ask the demonstrator.

Human Derived Products: All human derived products and equipment contaminated by the products such as blood and saliva must be considered as potential health hazards to you and to others in the laboratory.

• The student who has donated the product must be solely responsible for the handling of the product.

• All glassware and contaminated disposable material are to be disposed of as instructed by the demonstrator in charge.

Pre-lab Questions: Read and complete the following before entering the laboratory.

As you read the descriptions provided, label the parts on the diagram on the following page.

Ocular Lens

• Also known as the eyepiece • May be one only or a set of two • Lens closest to your eye, usually the highest part of the microscope • Includes a magnification factor engraved on the barrel, e.g. 10x indicates the

image is magnified ten times • Should be cleaned only with lens cleaner and lens paper

Nosepiece

Revolving circular mechanism that holds the different objective lenses Rotation of this changes the objective lenses

Objective Lenses

• Individual lenses attached to the nosepiece • Includes a magnification factor engraved on the barrel • Changed by rotating the nosepiece

Arm • Provides the safest way to hold a microscope. Use two hands.

Stage

• Also called the mechanical stage • A surface that supports the slide, with the help of stage clips

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Stage controls

Usually located on the top or side of the stage Moves the slide around the stage.

Condenser

• Located under the stage • Focuses the light through the hole in the stage and onto the specimen • Adjusts the quality and the amount of light passing through the specimen • May be raised or lowered with the condenser-adjustment knob

Iris Diaphragm

• Located under the condenser • Adjusts the intensity of light passing through the specimen • Use the iris diaphragm lever to open or close

Coarse focus adjustment knob

• Large knob located on the arm • Adjusts the distance between the stage and the objective lens, in large

increments • Used initially to bring the specimen into focus. It is dangerous to use this

knob when the objective lens is already near the slide • Should be turned very slowly to avoid breaking the slide

Fine focus adjustment knob.

• Small knob located inside the course adjustment knob • Adjusts in small increments • Typically used after the objective lens is already near the slide and the

specimen is almost in focus Lamp

• Small light source located under the condenser • May be turned on / off with switch on the base

Base Square or horseshoe–shaped support for the whole microscope Usually quite heavy to prevent tilting

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Activity 1 - Preparing a Wet Mount Slide

Materials for this activity

Paper towel Clean slide and cover slip Dropper bottle of methylene blue stain. Dropper bottle of physiological saline solution Flat toothpick / paddle pop stick. Microscope

• Collect a microscope. It should always be carried in an upright position. • Never place the microscope close to the edge of the bench. • Never touch the lens with your hands. • Use the revolving nosepiece to change objective lenses. • Always locate the object to be studied with the scanning-power objective in

position and then turn to the higher power objectives if a more detailed study is required.

• Do not use the coarse adjustment when the high power objective is in position. It is difficult to control the movement of this objective through small distances except by use of the fine adjustment knob.

• Never focus upward while looking through the eyepiece. Rather, look at the objective from the side to prevent moving the body tube too far and possibly damaging the objective lens to say nothing of the possibility of smashing the slide.

1. Place a drop of saline solution on the clean slide. Gently scrape the inner surface

of your cheek with the flat part of the toothpick, (Be careful not to draw blood. If blood is drawn, tell the instructor.) Transfer the cells from the toothpick to the saline solution by carefully stirring the end of the toothpick with the cheek cells in the saline (Figure a).

2. Place a small drop of the methylene blue stain directly on the saline/cell smear

(Figure b). Stir it again with the toothpick and immediately dispose of the toothpick in the autoclave bag.

3. Hold the cover slip by the sides to avoid getting fingerprints on it. Place one edge

of the cover slip next to one edge of the smear, and then lower it slowly into the smear (Figure c). By lowering it slowly you will avoid forming air bubbles. Absorb any excess fluid around the edges of the cover slip using paper towel.

(a) Stir cheek cells into saline solution b) Add drop of stain to the saline/cell smear (c) Lower cover slip onto smear

4. Place the cheek smear slide onto the stage. Focus with the scanning power lens

first. Scan for some individual cheek cells first. Notice that each cell has a distinctive dark centre called the nucleus. Draw one of these cells in results table on the following page. Be sure to include the proportion of the cell to the whole visual field, including details like the shape of the cell and nucleus and if the cytoplasm appears cloudy, clear or grainy.

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5. Return to the scanning-power lens, and scan for a cluster of cheek cells. Again look for a cluster that is separate from the others. Draw the cluster of cells in your results table.

Results:

Individual cheek cell:

Scanning power Low power High power

Grouped cheek cells:

Scanning power Low power High power

10

Questions:

1. What is the total magnification if the ocular lens is 10x and the objective lens is 100x? Show the calculations.

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2. Why should you scan a specimen under scanning power before using higher magnifications?

3. How is poor light quality affected by raising the condenser?

4. How is poor light quality affected by opening the iris diaphragm?

5. What is the shape of the individual cheek cells?

6. What does the thickness and the shape of the cells tell you about their function?

7. Was the nucleus centrally or peripherally located in the cells?

8. How do you think cheek cells connect to each other?

9. Why did you make the slide using physiological saline solution? Could you just use water instead?

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Activity 2 - Basal Metabolic Rate

Basal Metabolic Rate (BMR) is the minimum amount of energy required to sustain life and its calculation forms the basis of estimating daily energy requirements. The BMR does not account for any activity the individual may participate in during the day. It simply estimates the energy needed to sustain a heartbeat, breathing and normal body temperature with the body at rest (not sleep) at room temperature.

Calculate your own and the class averages for BMR using one of the following formulae:

Male:

BMR = 66 + (13.7 x weight) + (5 x height) – (6.8 x age)

Female: BMR = 655 + (9.6 x weight) + (1.7 x height) – (4.7 x age) For these equations, use weight in kilograms and height in centimetres.

Calculations:

My height = cm

My weight = kg

The class average for males is calories per day.

The class average for females is calories per day.

Questions:

1. Which was higher – male or female?

2. Explain the results.

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Results:

My BMR is calories per day.

Activity 3 – Cardiovascular Physiology

Many of the functions of the cardiovascular system can be measured using a few simple instruments. These give an indication of the condition of various structures.

In this activity you will investigate, measure and record the following:

1. heart sounds using stethoscopes

2. pulse rate from two superficial pulse points (three repetitions) by palpation and

timing with a watch

3. blood pressure (using a sphygmomanometer) and pulse (radial) (a) at rest sitting (b) after 3 minutes of intense activity (immediately and 5 minutes after you cease)

1. Heart Sounds

To hear the heart sounds you need to use a stethoscope. The earpieces need to be cleaned with alcohol swabs before use by each student. Once the alcohol has dried, place the ear pieces in your ears and the “bell” of the stethoscope on your partner’s thorax just to the medial side of the left nipple at the 5 th intercostal space. A characteristic “lup dup” sound can be heard with each beat of the heart, representing the closure of the atrio-ventricular and semi-lunar valves respectively.

2. Pulse Pulse rate can be elicited by palpating one of the major superficial pulse points on the body. The common carotid artery (neck), radial artery (in the wrist on the thumb side) and femoral artery (groin) are frequently used.

Take 3 readings of your partner’s pulse at two pulse points over a 1-minute period, and compare the averages.

3. Blood Pressure Blood pressure is measured in millimetres of mercury using a sphygmomanometer.

Firstly determine the blood pressure in (a) a relaxed sitting position.

Collect a stethoscope, a sphygmomanometer and some alcohol swabs from the front bench. Clean the earpieces as before and remove any excess air from the cuff of the sphygmomanometer.

Wrap the cuff of the sphygmomanometer around your partner’s upper arm just above the elbow. Find the arrow marked on the cuff and adjust the cuff so that the arrow points to the brachial artery. Secure the cuff using the Velcro wrap.

Palpate (find by touch) the brachial artery and place the “bell” of the stethoscope on the artery. Inflate the cuff to approx 160 mm pressure. Keeping the stethoscope in place, slowly release the pressure valve on the sphygmo and listen for the first “thudding” sounds of blood moving through the partially blocked (occluded) artery. This is the systolic pressure. Record this on the results table.

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Continue to release the pressure until the sound muffles or disappears and again record the result. This is the diastolic pressure. Repeat once.

Have your partner repeat the procedure to find your blood pressure. Blood pressure is recorded as the systolic pressure divided by the diastolic pressure.

Now determine the blood pressure and pulse of your partner and yourself after 3 minutes of intense activity (immediately and at 5 minutes).

Results:

1. Pulse rate:

Pulse point 1 Pulse point 2

Reading 1

Reading 2

Reading 3

Average

Question: How could you explain the difference between the average results for the two different pulse points? 2. Blood pressure:

After 3 minutes of activity At rest

Question: Why did the blood pressure readings change depending on your activity?

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0 min 5 min

Blood pressure

Pulse

Blood pressure

Pulse at rest (average from above)

of 10 When you are exercising your muscles need more oxygen. The heart then pumps more blood around your body with more powerful contractions which will make your blood pressure increase. Your systolic blood pressure increases during exercise as the cardiovascular system delivers more blood to the working muscles and your diastolic blood pressure stays roughly the same or decrease slightly.

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