biology 2.5 - studying cells - using microscopes

Why are specimens stained?

To stop light and electron beams from passing straight through transparent objects

How are specimens usually stained for light microscopes? What is eosin and methylene blue used to stain?

• The stains are usually dyes

• The stains are taken up more by some sections than other = contrast

• Eosin stains cytoplasm

• Methylene blue stains DNA

• More than one stain can be used at once

How are specimens usually stained for electron microscopes?

• Only done for TEMs as electron beams pass through the specimen

• Objects dipped in a heavy metal (e.g. lead)

• The metal ions scatter the electrons = contrast

How do you prepared a dry mount for a light microscope?

1. Thinly slice specimen as light needs to be able to pass through it

2. Use tweezers to put specimen in the middle of a clean slide

3. Put over slip on top

How do you prepared a wet mount for a light microscope?

1. Pipette a small drop of water into clean slide and put specimen on top using tweezers

2. Put coverslip upright on slide and slowly lower it to avoid air bubbles

3. Put a drop of stain next to one edge of the coverslip and a paper towel on the opposites edge to draw the stain under the coverslip

How do you prepared a squash slide for a light microscope?

1. Wet mount prepared

2. Coverslip pushed gently down with a lens and paper towel to avoid breaking coverslip

How do you prepared a smear slide for a light microscope?

1. Use the edge of a slide to smear the sample on a different slide to get an even coating

2. Put coverslip on top

How do you use a light microscope?

1. Clip slide to stage

2. Select the lowest power objective lens

3. Bring stage to just below objective lens using the coarse adjustment lens and then lower stage until the specimen is roughly in focus

4. Adjust focus with the fine adjustment knob until clear image seen

5. If higher magnification needed repeat process with a higher powered objective lens

What is the eyepiece graticule?

Fitted into the eyepiece and has number but no units

What is the stage micrometre?

It is a slide that is placed in the stage before the specimen and has units to it can give the eyepiece graticules’ divisions a value at a particular magnification

How do you use the eyepiece graticule and the stage micrometre?

1. Line up eyepiece graticules and stage micrometre so that at least one division is lined up (e.g. each division on the stage micrometre = 0.1mm

2. See how many eyepiece graticule divisions = 1 stage micrometre division at this magnification (e.g. 4.5)

3. Work out size of 1 eyepiece graticule division (0.1/4.5=0.0222mm)

4. Replace stage micrometre with the specimen and use the new scale on the eyepiece graticule to work out its length / width