Practical 1.

1
A: Calibrating an eyepiece graticule
B: Preparing a slide of onion epidermal cells

Safety
Wear eye protection.

Take care when using sharps.

Take care when using mains-operated microscopes with water or solutions.

Wash hands after handling biological material.

Apparatus and materials

• microscope • forceps
• eyepiece graticule • mounted needle
• stage micrometer • 2% iodine in potassium iodide solution
• sharp knife or scalpel • dropping pipette
• small onion (Allium cepa) • filter paper
• slides and cover slips • eye protection

Introduction
In this practical, you will:
• calibrate an eyepiece graticule
• make a temporary preparation of some onion epidermal cells
• stain the cells so that you can see structures within them
• set up a light microscope and use it to make observations and measurements
• make a drawing of the onion cells
• use the eyepiece graticule you calibrated to measure the size of the cells.

Procedure

A Calibrating an eyepiece graticule

1 If it does not already have one, insert a graticule into the eyepiece of the microscope by
unscrewing the top lens, resting the graticule on the rim halfway down and replacing the top lens.
2 Place a stage micrometer slide on the stage of the microscope. The smallest division on the stage
micrometer equals 100 m.
3 Using the low-power objective, focus the microscope on the stage micrometer. Rotate the eyepiece
and move the slide to superimpose the scales of the eyepiece graticule and the stage micrometer
(shown in the diagram on the next page).

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4 Count the number of divisions on the eyepiece graticule equivalent to 100  m on the stage
micrometer and then calculate the length that one eyepiece division is equivalent to. For example,
if three divisions are equal to 100 m, then each division is equal to 33.3 m at low power. Record
your answer.
5 Repeat step 4 for the medium-power and high-power objectives. You have now calibrated the
eyepiece graticule and you can use it to measure cells in the preparation below, or in the
preparations in the next practicals.

B  Preparing a slide of onion epidermal cells

1 The fleshy layers inside an onion are known as ‘leaves’ and store nutrients. Cut open an onion and
separate some of the leaves. Peel off one of the thin layers of epidermal tissue from the inner
concave surface of a leaf and transfer it to a drop of water on a microscope slide. Use forceps and
a mounted needle to make sure that the tissue is not folded.
2 Place two drops of iodine solution onto the tissue. Gently lower a coverslip onto the slide using a
mounted needle. Use a piece of filter paper to absorb excess stain. Place some filter paper over the
coverslip and gently press to flatten the specimen.
3 Place the slide on the stage of the microscope and use the low-power objective to locate the cells.
Now use the high-power objective to select three adjacent cells that are clearly visible in your field
of view.
4 Make a large, labelled drawing of these three epidermal cells. Use a sharp pencil (HB) and a ruler
to draw the label lines and labels.
5 Use the eyepiece graticule to measure the length of one of the epidermal cells that you have drawn.
Now measure the same cell in your drawing.
6 Calculate the magnification of your drawing, using the formula:

Remember that both lengths must be measured in the same units, e.g. micrometres (m). Write the
magnification underneath your drawing.

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