Practical Zero

Question 1

Visualising materials in tissue sections:

What technique uses labelled antibodies to locate specific proteins or other antigens?

What technique uses organic dyes to stain tissues and cell structures like nuclei and cytoplasm?

Answer 1

Immunohistochemistry

Histology

Question 2

Name the 2 types of electron microscopes.

Answer 2

1. Transmission

2. Scanning

Question 3

Name a fixative used in the preparation of samples for microscopy.

Answer 3

Gluraraldehyde

Question 4

What is the limiting factor in the resolution of detail by light microscope?

Answer 4

The wavelength of the light!

Question 5

What function does ethanol play in the processing of material for microscopy?

Answer 5

Removal of water

Question 6

What does Eosin stain bind to?

What does PAS staining bind?

What does Haematoxylin bind?

Answer 6

Protein

Complex carbohydrates

Nucleic Acids (DNA)

Question 7

If you observe a specimen using a light microscope with a 10x objective lens, what is the final magnification?

Answer 7

100

Question 8

In immunohistochemistry the indirect method provides a HIGHER fluorescent signal than the direct method. True or false?

Answer 8

True!

Question 9

With the electron microscope, you can observe LIVE cells and tissues. True or false?

Answer 9

FALSE

Question 10

What is the bottom part of the microscope called?

What is the part above the Illuminator called, that is split into 2 parts?

Answer 10

Illuminator

Condenser lenses

Question 11

What is meant by the term homogenisation?

Answer 11

A mechanical OR non-mechanical process by which cells and tissues are broken into fragments small enough to create a uniform, stable emulsion.

Question 12

Why do we use Iodonitrotetrazolium (INT) in the reactions?

** Sub-cellular fractionation practical - 0 **

Answer 12

INT is an artificial electron acceptor!

INT makes a formazan red dye that can be viewed down the microscope!

Question 13

What should be done when preparing samples for centrifugation?

Answer 13

Samples should have the SAME volume in, measured by weight or pipettes volume.
Samples should be distributed EVENLY around the centrifuge!
Samples should be labelled to identify the contents and the owner.

Question 14

Practical 0 - sub cellular fractionation

Absorbic acid is better know as…

What property of absorbic acid makes it useful in this experiment?

Why is absorbic acid used as a control?

Answer 14

Vitamin C

It can donate electrons to INT, changing its colour to purple!

It proves that the experiment works because INT should turn oriole when it is reduced! It proves that the homogenate has mitochondria in it.

Question 15

What is the CELLULAR function of succinate dehydrogenase (SDH)?

Answer 15

To convert succinate to fumerate in the Krebs (citric acid) cycle.

Question 16

Practical 0 - sub cellular fractionation

What purple staining indicate?

Answer 16

Purple staining = fragment contains mitochondria = as there is presence of the enzyme succinate dehydrogenase which resides in the mitochondrial matrix!

Question 17

Practical 0 - sub cellular fractionation

Which set of samples were the darkest?

Answer 17

Those with the succinate added as the substrate!

Question 18

Practical 0 - sub cellular fractionation

Why would you need to compare your INT colour changes with what you observed under the microscope?

Answer 18

To see how pure the samples were, whether there was any cross contamination between fractions.

To help us explain why we might have got these results from the INT reactions.

To identify if the nucleus has enzymes that release electrons to allow reduction reactions.

Question 19

Practical 0 - sub cellular fractionation

Why did you get a deeper colour change in the fraction containing the mitochondria?

Answer 19

This was where succinate dehydrogenase was working and producing electrons to donate to INT, inducing the colour change.