PAG 1 Microscopy

Question 1

Explain how to calibrate an eyepiece graticule (5)

Answer 1

1. Set the microscope to the required magnification for viewing the sample.
2. Place a stage graticule (stage micrometer) on the stage.
3. Align the scales of the stage and eyepiece graticules as shown in a diagram.
4. Count the number of eyepiece divisions that match one stage micrometer division.
5. Use the known value of each stage division to calculate the length of one eyepiece division

Question 2

State what must be done if the microscope magnification is changed (2)

Answer 2

• The eyepiece graticule must be recalibrated for each objective lens.
• Creating a calibration factor for each lens.

Question 3

Define staining and its purpose (2)

Answer 3

• Staining involves applying a dye to a specimen to improve contrast and make structures more visible.
• Differential staining uses multiple dyes that bind to specific cell structures, colouring each differently for easier identification

Question 4

State five examples of stains and what they detect (5)

Answer 4

• Acetic orcein: binds to DNA, staining chromosomes dark red.
• Eosin: stains cytoplasm dark red or pink.
• Iodine: stains starch blue-black (violet under a microscope).
• Iodine in potassium iodide solution: stains cellulose yellow.
• Haematoxylin: stains RNA/DNA purple or blue, and methylene blue: stains DNA blue.

Question 5

Describe the wet mount slide preparation method (4)

Answer 5

1. Add a drop of water to the slide using a pipette.
2. Place the specimen in the water with tweezers.
3. Position the cover slip upright next to the droplet, then gently lower it to avoid air bubbles.
4. If needed, place stain at one edge of the cover slip and draw it through using a paper towel on the opposite edge.

Question 6

Describe the dry mount slide preparation method (3)

Answer 6

1. Cut the specimen into a thin slice so light can pass through.
2. Place it in the middle of the slide using tweezers.
3. Put a cover slip on top

Question 7

Explain how to use a light microscope (5)

Answer 7

1. Clip the prepared slide onto the stage.
2. Select the lowest-powered objective lens.
3. Use the coarse adjustment knob to bring the lens close to the slide without touching it.
4. Look through the eyepiece and focus by moving the lens away from the slide using the fine adjustment knob.
5. Switch to a higher-powered lens if needed and refocus.

Question 8

Describe how to measure specimen size with a microscope (3)

Answer 8

1. Use an eyepiece graticule that has been calibrated.
2. Count the number of eyepiece divisions across the structure.
3. Multiply the number of divisions by the value of one division to find the actual size.

Question 9

Describe how to measure the diameter of an irregularly shaped object (1)

Answer 9

Measure the greatest distance from one side to the other

Question 10

State the rules of producing biological drawings (6)

Answer 10

• Do not use shading; label shaded areas instead.
• The drawing should fill at least half of the page.
• Label lines must be horizontal, drawn with a ruler, and must touch the structure without overlapping other lines.
• Drawing lines should be continuous and not feathery.
• Include a scale, e.g. based on the magnification of the image.
• Draw in pencil, ensuring it resembles the actual specimen.