Microscopy required practical

Aim: To prepare a stained slide of plant or animal cells, view it with a light microscope and produce a labelled drawing with a calculated magnification.

Variables

• Independent: Not a comparative investigation; the type of cell viewed (e.g. onion epidermis or cheek cells) and the objective lens used can be varied.
• Dependent: The detail visible and the size of the cells or structures observed.
• Control: Thickness of the specimen (a single layer of cells); Amount of stain used; Lighting setting of the microscope when comparing specimens

Equipment

• Light microscope with lamp or mirror
• Glass microscope slides and cover slips
• Onion and a white tile
• Scalpel and forceps
• Iodine solution in a dropping bottle (or methylene blue for cheek cells)
• Mounted needle
• Paper towel
• Ruler, or eyepiece graticule and stage micrometer

Method

1. Put a drop of water in the centre of a clean slide.
2. Peel a thin, see-through layer of epidermis from the inside of an onion scale leaf using forceps.
3. Lay the epidermis flat on the water drop, avoiding folds.
4. Add one drop of iodine solution to stain the cells so the nucleus and cell wall show up.
5. Lower a cover slip at an angle onto the specimen using a mounted needle, so that no air bubbles are trapped; blot off extra liquid.
6. Place the slide on the stage and select the lowest power objective lens.
7. Move the stage close to the lens, then use the coarse focusing knob to move it away until the cells come into focus.
8. Switch to a higher power objective lens and use the fine focusing knob to sharpen the image.
9. Draw a few cells with a sharp pencil using clear, unshaded lines, label the visible parts and record the total magnification (eyepiece × objective).

Safety

• Iodine solution is an irritant and stains skin; wear eye protection and wipe up spills.
• Take care with the scalpel and handle slides and cover slips gently because broken glass is sharp.
• If the microscope uses a mirror, never point it at direct sunlight as this can damage the eyes.

Results and calculations

Produce a labelled pencil drawing, noting the magnification used. To find the real size of a cell use magnification = image size ÷ real size, rearranged to real size = image size ÷ magnification, making sure both sizes are in the same units (1 mm = 1000 µm).

Common mistakes and improvements

• Trapping air bubbles under the cover slip; lower it slowly at an angle from one edge.
• Using a specimen that is too thick so light cannot pass through; peel a single layer of cells.
• Starting on high power and crashing the lens into the slide; always begin on low power and focus by moving the stage away from the lens.
• Mixing units when calculating size; convert everything to micrometres or millimetres before dividing.

Exam tips

• Know why each step is done: stain makes structures visible, the cover slip keeps the specimen flat and stops it drying out, and a thin layer lets light through.
• Be ready to rearrange magnification = image size ÷ real size and convert between mm and µm, giving answers in standard form where asked.
• Drawing marks come from smooth continuous lines, no shading, labels with ruled lines that touch the structure, and a title with magnification.
• Explain why chloroplasts are not seen in onion epidermis cells (they are not exposed to light) and why a light microscope cannot show ribosomes (limited resolution).
• Compare light and electron microscopes in terms of magnification and resolution.