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Lecture 2 Flashcards

Tissue prep and staining (50 cards)

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1
Q

What are the five main components of the compound light microscope?

A
  • light source
  • condenser
  • stage
  • objective lens
  • ocular lens
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2
Q

What is the pro of the compound light microscope?

A

The ability to resolve structural detail

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3
Q

List the cons of a compound light microscope

A
  • little magnification ability
  • the specimen must be thin
  • in an unstained specimen there is little contrast`
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4
Q

How does a phase contrast microscope work?

A

When light passes through a specimen it converts phase shifts that are invisible to the eye into brightness changes that the eye can see

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5
Q

What are the advantages of using a phase contrast microscope?

A
  • this microscope is useful when looking at unstained specimens
  • it can be useful when examining living cells
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6
Q

When is a fluorescence microscope used?

A
  • to detect antigens or antibodies in immunochemical staining
  • detect fluorescent tracers that have been injected into cells
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7
Q

Describe the function of a confocal scanning microscope

A

Laser beams are moved across the surface of a specimen where the data is recorded and translated into a 3D image and displayed on a monitor

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8
Q

Describe the advantages of a confocal scanning microscope

A
  • thin optical images
  • the computer program retracts any out-of-focus images
  • 3D reconstructions can be made by stacking images
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9
Q

In transmission electron microscopy (TEM) what is used instead of light?

A

A beam of electrons

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10
Q

What are the four steps to prepare tissue for observation?

A
  • Fixing
  • Dehydration
  • Clearing alcohol
  • Embedding
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11
Q

Why must tissues be fixed?

A

The tissue will keep deteriorating until the tissue is fixed in a fixitive. The fixitive will also help the tissue harden.

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12
Q

What is one of the most widely used fixing agents?

A

10% buffered formalin

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13
Q

What is a downside of fixatives?

A

Radically distorts the specimen

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14
Q

What are acidic fixatives best used for?

A

When you are wanting to fix the chromatin, nucleoli, and spindle fibers. It will NOT fix mitochondria or nucleoplasm

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15
Q

List three types of acidic fixatives

A
  • carnoy’s fluid
  • zenker’s fluid
  • bouin’s fluid
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16
Q

Fixative good for preserving glyogen in animal tissues is

A

Carno’ys fluid

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17
Q

An acidic fixative that must be washed carefully to avoid formation of black crystals is

A

Zenker’s fluid

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18
Q

Fixitive that gives great cytological detail, but needs thorough washing is called:

A

Bouin’s Fluid

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19
Q

If you are looking to fix mitochondrial structures what kind of stain would you want to use?

A

A basic fixative

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20
Q

In basic fixatives the chromatin is dissolved, what is an example of a basic fixative?

A

Zirkle-Erliki fixative

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21
Q

List the two fixatives ideal for TEM

Study These Flashcards
A
  • Glutaraldehyde

- Osmium Tetroxide

22
Q

Why must tissue be dehydrated prior to embedding?

Study These Flashcards
A

Because the tissue will be embedding in a hydrophobic material all the water must GTFO

23
Q

How is tissue dehydrated?

Study These Flashcards
A

Tissue is placed in increasing strengths of ethanol until all the water is removed.

24
Q

When you need to remove the ethanol from the tissue what substances are typically used?

Study These Flashcards
A

Xylene or cedar oil

25
What is different about the embedding process for TEM?
Instead of paraffin, the tissue is infiltrated with a monomeric resin that is then polymerized.
26
What is an advantage of using a rotary microtome over a hand-held microtome?
The rotary microtomes advance at a fixed distance with each rotation, which makes sectioning uniform and easier.
27
When sectioning for TEM how thick should the sections be?
50 - 150 nm thick
28
Why must the sections for TEM be floated onto the plastic-coated copper grid?
The sections are too fragile for normal handling
29
Why should tissues be stained prior to observation?
Since animal tissues are typically colorless, the stains color them artificially to bring out detail
30
What is the most common stain used for routine staining
Routine H&E staining (Hematoxylin and Eosin)
31
If you wanted to look at elastic material like connective tissue what stain would you use?
Orcein, and resorcin fuchsin stains
32
What is silver impregnation staining used for?
Used to show reticular fibers and basement membranes
33
What is an example of a fat-soluble stain?
Sudan
34
Basic dyes will react with what kind of groups of tissue components?
The anionic groups: - phosphate - sulfate - carboxyl
35
List four examples of basic dyes
- methylene blue - toluidine blue - methyl green - pyronine G
36
What kind of linkages do acidic dyes form with cationic groups?
Electrostatic linkages
37
List four examples of acidic dyes:
- acid fuchsin - eosin - orange G - aniline blue
38
Define metachromasia
Metachromasia refers to the phenomenon where a dye will change color AFTER it has reacted with components in the tissue. Ex: Toluidine Blue
39
Staining for TEM requires what category of stains?
Ions of heavy metals
40
Explain what histochemistry is
Histochemistry techniques can be used to study the chemistry of tissues and cells
41
Frozen sections are best used to observe:
Lipids
42
What does a Schiff reagent reaction depend on?
the formation of aldehyde groups
43
What is the Fuelgen reaction?
The fuelgen reaction is a type of Schiffs where HCl exposes aldehyde groups on deoxyribose
44
What is the Periodic acid-Schiff reaction (PAS)?
Aldehyde groups are formed by the cleaving of bonds of adjacent carbons on carbohydrates by periodic acid.
45
What is the advantage of immunocytochemical staining techniques?
By using monoclonal antibodies you can look for the specific presence of antigens
46
Give three examples of antigens
- proteins - glyoproteins - proteoglycans
47
What does the term "monoclonal" refer to?
A single immune response to an antigen
48
What are three ways to directly label antibodies?
- fluorescent dye - visible substance - gold or ferritin
49
Polyclonal antibodies have what characteristic?
Multiple binding sites that generate a variety of different antibodies
50
A second antibody is used during what kind of labeling?
Indirect labeling

Histology (12 decks)

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