Histology Exam 1: Slide Preparation

Front 1

Why fix a slide?

Back 1

1) Terminates Cell Metabolism
2) Prevents enzyme degredation of cells by a process called autolysis
3) Kills pathogenic viruses, bacteria, and fungi
4) Hardens tissue by cross-linking or denaturing proteins

Front 2

3 Steps in Slide Preparation

Back 2

1) Fixation
2) Infiltration with paraffin
3) Staining

Front 3

Why should you use a neutral, buffered formalin solution in fixation?

Back 3

The neutral pH prevents the formation of acid hematin crystals in the tissue when formalin reacts with hemoglobin in an acidic environment

Front 4

How does formaldehyde act as a preservative and why is it preferred?

Back 4

It preserves cell structure and extracellular components by reacting with amino groups of protein, resulting in a cross-linking between lysine residues. It minimally alters the 3D structure of most proteins, preserving their immunogenicitiy and allowing subsequent use of labeled antibodies in immunofluorescent techniques and immunohistochemistry.

Front 5

Why are slide tissues infiltrated with paraffin?

Back 5

To keep the tissue rigid while it is sliced into extremely thin sections.

Front 6

How do you infiltrate a slide with paraffin?

Back 6

The slide is washed until free of fixative, then dehydrated in a series of increasingly concentrated alcohols (ending in 100%). It is finally placed in a clearing solution that consists of organic solvent (e.g. xylene or toluol) and immersed in liquid paraffin at 57-59 degree C. The cooled, hardened tissue in a paraffin block is mounted and sectioned.

Front 7

What is an unstained paraffin block?

Back 7

A specimen after paraffin infiltration (and before staining) sectioned off from a paraffin block.

Front 8

What do H&E stand for and which stain is water-soluble?

Back 8

H= hematoxylin (water soluble)
E= eosin (alcohol soluble)

Front 9

Following paraffin filtration, what must be done to the slide before it is "ready"?

Back 9

First paraffin must be dissolved from the tissue with xylol or toluol. The tissue is rehydrated with decreasing alcohol solutions (ending in water). After staining, tissue is dehydrated a second time, re-immersed in xylol or toluol, and covered with a thin glass cover slip held in place by non-aqueous mounting medium.

Front 10

What histologic structures *cannot* be seen on an H&E slide?

Back 10

Basement membranes, intra-cellular lipid, elastic connective tissue, and reticular fibers.

Front 11

Name the histologic structure(s) we look for using Periodic acid Schiff (PAS) stain.

Back 11

Basement membrane (purple), Glycogen (magenta)

Front 12

Name the histologic structure we look for using Masson's or Mallory's stain.

Back 12

Muscle (red)

Front 13

Name the histologic structure we look for using trichrome stain.

Back 13

Collagen (blue)

Front 14

Name the histologic structure we look for using Weigert's stain.

Back 14

Elastic fibers (black)

Front 15

Name the histologic structure we look for using silver stain.

Back 15

Reticular fibers (black)

Front 16

Name the histologic structure we look for using Oil and red O stain.

Back 16

Lipid (red)

Front 17

Name the histologic structure we look for using Wright's or Wright-Giemsa stain.

Back 17

Blood cells, cytology

Front 18

Name the histologic structure we look for using Toluidine blue (T-blue) stain.

Back 18

Mast cell granules (metachromasia)

Front 19

What are common sources of artifacts in prepared slides?

Back 19

Insufficient/prolonged tissue fixation (prolonged is especially common when not using formalin), failure to completely dehydrate the tissue prior to paraffin embedding, and use of paraffin that is too hot during the embedding process.

Front 20

Differentiate between polyclonal and monoclonal antibodies.

Back 20

Polyclonal antibodies are produced in an immunized animal while monoclonal antibodies are produced in a continuously replicating antibody-producing cell line.

Front 21

What color does fluorescein appear when viewed under UV light?

Back 21

Bright apple green