Using a Light Microscope

Main parts of the microscope

• Eyepiece lens, the lens closest to the eye, usually with a magnification of x5 or x10.
• Objective lenses, a set of lenses on a rotating nosepiece, typically low power (x10), medium power (x40), and high power (x100).
• Stage, the flat platform where the slide is placed and held by clips.
• Diaphragm, controls the amount of light passing through the specimen from the light source below.
• Coarse adjustment knob, moves the stage or lens tube a large distance to bring the specimen roughly into focus.
• Fine adjustment knob, moves the stage or lens tube a small distance to sharpen the focus.
• Body tube and base, the body tube connects the eyepiece to the objective lenses, and the base supports the whole microscope.

Steps to focus a specimen

1. Place the prepared slide on the stage and secure it with the clips, positioning the specimen over the hole in the stage.
2. Rotate the nosepiece so that the low-power objective lens is directly above the specimen.
3. Looking from the side, use the coarse adjustment knob to lower the objective lens close to the slide without touching it.
4. Looking through the eyepiece, slowly turn the coarse adjustment knob to raise the lens tube until the specimen comes roughly into view.
5. Use the fine adjustment knob to bring the specimen into sharp focus.
6. Adjust the diaphragm if needed so the specimen is clearly lit but not too bright.

Moving from low power to high power

Always focus a specimen under low power first, since it has a wider field of view and makes the specimen easier to locate. Once the specimen is centred and in focus, rotate the nosepiece to the next higher-power objective lens.

Because most microscopes are parfocal, the specimen should stay roughly in focus, so only the fine adjustment knob is needed to sharpen the image again. The coarse adjustment knob should not be used at high power, as it can move the lens into the slide and break it.

Magnification and calculating image size

The total magnification of a specimen is the magnification of the eyepiece lens multiplied by the magnification of the objective lens in use. For example, a x10 eyepiece with a x40 objective gives a total magnification of x400.

To work out the real size of a structure, measure its size in the image and divide by the magnification: actual size = image size ÷ magnification. If a cell measures 40 mm across in a drawing made at x400, its real width is 40 ÷ 400 = 0.1 mm, or 100 micrometres.

Keep the image size and the actual size in the same unit before dividing.

Safety when using a microscope

• Carry the microscope with one hand under the base and one hand on the arm, and set it down gently on a flat bench.
• Handle glass slides and cover slips by their edges, as they are thin and break easily; clear away any broken glass at once.
• Take care with the lamp or mirror and the power lead, as the lamp can become warm and the electrical parts should stay dry.
• Clean the lenses only with lens tissue, not with fingers or ordinary cloth, to avoid scratching them.

Paper 3-style questions

Question 1, procedure. A student cannot find the specimen when looking through the eyepiece. Suggest the correct order of steps to locate and focus it.

Model answer. Start with the low-power objective in place. Looking from the side, use the coarse adjustment knob to bring the lens close to the slide.

Then, looking through the eyepiece, turn the coarse knob to move the lens away until the specimen appears, and finish with the fine adjustment knob for a sharp image before switching to higher power.

Question 2, calculation. A specimen is viewed with a x10 eyepiece and a x40 objective. State the total magnification and calculate the actual length of a cell that appears 20 mm long.

Model answer. Total magnification = 10 × 40 = x400. Actual length = image length ÷ magnification = 20 ÷ 400 = 0.05 mm, which is 50 micrometres.

Question 3, inference. When the student switches to high power the field of view becomes darker and shows fewer cells. Explain both observations.

Model answer. The field of view is smaller at high power, so fewer cells are seen but each appears larger. It looks darker because the higher-power lens covers a smaller area and lets less light through, so the diaphragm should be opened wider to brighten the image.

Common mistakes

Source:SRC-DSKP-EN

Frequently asked questions