Investigative Techniques

Question 1

What are 3 requirements needed to image tissues by light microscopy?

Answer 1

• preserve the tissue
• embed the tissue, allowing it to be spliced
• stain the tissue

Question 2

What substance is normally used to preserve tissue in light microscopy?

Answer 2

Formalin

Question 3

What substance is the tissue usually embedded in to allow it to be cut?

Answer 3

Melted paraffin

Question 4

What is normally used to stain tissues used in light microscopy?

Answer 4

Haematoxylin and eosin

Question 5

What structure does haematoxylin stain? What colour does it stain?

Answer 5

Haematoxylin stain the nucleus blue

Question 6

What structure does eosin stain? What colour does it stain?

Answer 6

Eosin stains the cytoplasm and extracellular matrix pink

Question 7

Name a similarity between indirect immunohistochemistry and immunofluoresence.

Answer 7

They both require binding the of a primary antibody

Question 8

How does indirect immunohistochemistry produce a signal that can be read?

Answer 8

A secondary antibody attaches to the primary antibody - the secondary antibody has an attached enzyme which converts its substrate to product - this product will give off a signal and show Ho much antibody (and as such how much protein the antibody is attached to) is present

Question 9

How does immunofluoresence produce a signal that can be read?

Answer 9

The primary antibody will have an attached fluorescent tag, which will emit a fluorescent signal if the antibody attaches to its target protein

Question 10

What is the difference between high and low frequency, in regards to their wavelengths and resolution?

Answer 10

Low frequency has longer wavelengths but poorer resolution

High frequency has shorter wavelengths but greater resolution

Question 11

How does the resolution of a light microscope compare to an electron microscope?

Answer 11

The resolution of a light microscope is much worse - the wavelength of a light microscope is 400um, while the wavelength of a electron microscope is 1um - electron microscopes therefore have a resolution that is 400x more powerful

Question 12

Compare the magnification power of a light microscope and an electron microscope.

Answer 12

Magnification in electron microscope is around 250,000x, while magnification in light microscopes is around 1,000x - electron microscopes are therefore 250x greater magnification power than a light microscope