2 - Observe samples using a light microscope

What is the function of a stage micrometer?

To calibrate an eyepiece graticule, so that it can be used to make measurements

How is an eyepiece graticule calibrated?

1. Select the desired magnification & objective lens & place the stage micrometer on the stage

2. Line up the scales of the stage micrometer & the eyepiece graticule

3. Count the number of divisions on the eyepiece graticule equivalent to each division on the stage micrometer

4. Calculate the length of one division of the eyepiece graticule

What equipment is required for the practical?

• Plant stem & light microscope

• Eyepiece graticule & stage micrometer

• Toluidine Blue O stain

• Microscope slides/coverslips

• Knife & paintbrush

• White tile & watch glass

• Pipette & lens tissue

• Mounted needle & paper towels

What is the method for this practical?

1. Calibrate the eyepiece graticule

2. Cut thin transverse sections of a plant stem on a white tile using a wet knife to reduce friction & select the two thinnest sections

3. Mount one section in water on a microscope slide & lower the cover slip down carefully, ensuring there are no air bubbles

4. Add toluidine blue O stain (adds contrast) to the other section & leave for 3 mins, then mount in water on another microscope slide & add a cover slip

5. Place the slide under a microscope & set the objective lens on the lowest magnification, then use the coarse adjustment knob to move the lens down to just above slide

6. Use the fine adjustment knob to re-adjust the focus until the image is clear

7. Observe & draw the stem

8. Measure the size of stem diameter & vascular bundle

What are the requirements for label lines in biological drawings?

Lines must be horizontal, straight (drawn with a ruler) & must not overlap

What is the risk, safety precaution & in emergency for the hazard: toluidine blue O stain

Risk:

• may cause harm/irritation to eyes or in cuts

Safety precaution:

• keep away from naked flames

• wear eye protection

• avoid contact with skin

In emergency:

• put out fire & seek assistance

• wash off skin immediately

• flood eye/cuts with cold water

What is the risk, safety precaution & in emergency for the hazard: knife

Risk:

• cuts from sharp object

Safety precaution:

• cut away from fingers

• use forceps to hold sample whilst cutting

• keep away from edge of desk

In emergency:

• elevate cuts & apply pressure

• seek medical assistance

What is the risk, safety precaution & in emergency for the hazard: broken glass

Risk:

• cuts from sharp object

Safety precaution:

• take care when handling slides & coverslips

• keep glassware away from the edge of the desk

In emergency:

• elevate cuts & apply pressure

• don’t remove glass from wound

• seek medical assistance

Outline the procedure to prepare a slide

1. Stain the sample with an appropriate dye

2. Mount the sample on the slide

3. Place a cover slip carefully on the slide, avoiding air bubbles

What is the formula for magnification?

Magnification = image size / actual size