Core Practical 5: (i) Use a light microscope to make observations

and labelled drawings of suitable animal cells;(ii) Use a graticule

with a microscope to make measurements and understand the
concept of scale

Objectives
● To be competent in the use of a microscope at high and low power, including the use of a
graticule (eyepiece micrometer) to make measurements
● To know how to record observations using appropriate biological drawings
● To understand the importance of staining specimens in microscopy
Safety
● Methylene blue may be harmful if swallowed but is not otherwise classified as hazardous.
However, there is always the possibility of unknown effects, so stains should be used with
caution.
● Avoid skin contact with iodine and methylene blue stains. Wear gloves and eye protection when
handling the stain. Clean up any spills immediately.
● Make sure all biohazardous material is placed in disinfectant solution as soon as possible.
Cotton buds should only be used once, by one individual.
● If you are using a microscope with daylight illumination (a mirror) do not place it where sunlight
might strike the mirror as this will damage your retina and may cause blindness.
Maths skills
● Use ratios, fractions and percentages.
● Change the subject of an equation.
● Make order of magnitude calculations.
Equipment
● eye protection
● cotton buds
● microscope with eyepiece graticule
● stage micrometer slide
● methylene blue
● glass microscope slides
● coverslip
● dropping pipette
● lens tissue
● absorbent paper (e.g. paper towel)
● large beaker of disinfectant solution

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Practical activities have been safety checked but not trialled. Users may need to adapt the risk assessment
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 Diagrams

figure A Calibrating the stage micrometer using an eyepiece graticule

figure B Section of an animal cell as seen through a microscope

Procedure
Procedure – Part 1: Calibration
First, you will need to calibrate the eyepiece graticule (see figure A).
1. Place a micrometer slide on the stage of the microscope and focus on the micrometer scale,
using the low-power objective. The smallest division of the micrometer scale is usually 100 µm.
2. Move the slide and rotate the eyepiece to align the scales of the eyepiece graticule and the
stage micrometer in the field of view.
3. Count the number of divisions (eyepiece units or epu) on the eyepiece graticule that are
equivalent to a known length on the micrometer slide and work out the length of one eyepiece
unit. For example, if 100 µm is

4. Repeat steps 1–3 with the medium- and high-power objectives.

Procedure – Part 2: Making observations
1. Wash your hands with soap and water.
2. Take a cotton bud and gently rub it on the inside of your cheek, then rub the cotton bud in a
small circle in the centre of a glass slide. Immediately place the cotton bud in a beaker of
disinfectant solution.

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Practical activities have been safety checked but not trialled. Users may need to adapt the risk assessment
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 3. Add a few drops of methylene blue to the sample, then cover with a cover slip.
4. Turn the objective lens to low power and examine the stained slide under the microscope. To
do this, bring the lens as close to the slide as possible while watching it from the side of the
instrument. Then, looking through the eyepiece, use the coarse focusing knob to focus, moving
the lens away from the stage. This avoids damage to the slide or lens. Finally, use the fine
focus until a clear view of the cells is established.
5. Carefully sketch a few of the cells. Use figure B to help you identify the parts of the cell.
6. Use the eyepiece graticule to measure a cell’s diameter. Add a scale bar to your diagram. Add
a title and include the magnification at which you made your observations. For example, with an
eyepiece lens magnification of ×10 and an objective of ×10, the total magnification will be ×100.
Remember, this is not the same as the magnification of the drawing.
7. Now, turn the objective disc to the medium-power lens, and focus until the cells are clear and
distinct. Identify as many details of the cells as you can.
8. Finally, turn the objective disc to the high-power lens and focus using the fine-focusing knob
only. Draw and label the detail of the cells as accurately as you can.
9. Measure the length and breadth of two cells. Record these measurements in your diagram.
10. Place the glass slide in the beaker of disinfectant solution.
Learning tips
● To be able to answer examination questions about the magnification of images, make sure you
learn the following equation:

● You should know how to rearrange the magnification formula to calculate any of the values.
Remember to convert all lengths to the same units, usually µm.

Tips for good biological drawing

● Use a sharp HB pencil. Keep lines clear and continuous, not feathery or sketched.
● Draw only what you see. Do not draw stylised patterns, and do not make it up.
● Start with an outline. Keep it large and think about proportions.
● Do not use shading or colour.
● Draw label lines in pencil with a ruler. Lines should not have arrowheads and should just touch
the item to be labelled.
Questions
1. Use the scale bar on your diagram to calculate the magnification of the image you have drawn.
2. Explain briefly how cells can be measured using a microscope.
3. Suggest improvements to the method that was used to describe the size of cells in a tissue.

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Practical activities have been safety checked but not trialled. Users may need to adapt the risk assessment
information to local circumstances. 2
 Exam-style questions
1. Under a high-power objective lens, 300 µm on a stage micrometer was found
to be equivalent to 87 eyepiece units.
(c) What is the size of each eyepiece unit in µm?
(1)
(d) Using the same objective, the diameter of a red blood cell was measured as
2.5 eyepiece units. What is the actual diameter of the red blood cell?
(2)
2. A cell from epithelial tissue in the trachea was found to be approximately cylindrical in shape.
The cell had a length of 20 µm and a diameter of 4 µm.

The volume (v) of a cylinder may be calculated from v = πr2h.

Calculate the volume of the cell. Show your working.

(3)
3. State the purpose of staining specimens in light microscopy.
(4)

© Pearson Education Ltd 2018. Copying permitted for purchasing institution only. This material is not copyright free.
Practical activities have been safety checked but not trialled. Users may need to adapt the risk assessment
information to local circumstances. 3