Chapter 1 - Histology and its method of study

Question 1

Microanatomy

Answer 1

• structure of individual cells w/n a tissue, and structure of different types of tissues
• function of cells and tissues, roles in different organ systems

Question 2

Who were the scientists that received a Nobel prize for histology? What did they do?

Answer 2

• Santiago Ramon y Cajal and Camillo Golgi
• determined the structure of neurons in the brain with staining method

Question 3

What is the ideal goal of specimen prep?

Answer 3

to preserve the tissue’s structure
- sometimes not possible because of where the sample comes from or other circumstances

Question 4

Fixation

Answer 4

• Step 1 of specimen prep
• strong chemical used to preserve structure and prevent microbial degradation
• irreversibly cross links and denature proteins

Question 5

What fixatives are used for LM and EM?

Answer 5

• 4% formaldehyde for LM
• Glutaraldehyde and OsO4 for EM
• both methods prevent immunochemistrical methods

Question 6

Dehydration and clearing

Answer 6

• Step 2 of specimen prep
• water from fixation is removed using dehydrator like alcohol, and then the dehydrator is cleared

Question 7

What are the chemicals used for dehydration and clearing?

Answer 7

• ethanol - dehydration
• xylene - clearing

Question 8

Embedding

Answer 8

• Step 3 of specimen prep.
• Sample is put into some kind of wax or solid matrix, which is then trimmed away so the sample is ready for sectioning and staining

Question 9

What different matrixes are used for embedding?

Answer 9

• Parffin - LM
• Epoxy resin for EM, much more stable then paraffin b/c EM requires much thinner slices

Question 10

Sectioning, what are the different types for LM and EM?

Answer 10

• Step 4 of specimen prep
• Microtome - glass knife, used for LM, 1-10 micrometers
• Ultramicrotome - diamond knife, used for EM, 50 nanometers

Question 11

What type of sectioning does not require fixation?

Answer 11

• cryosectioning
• frozen w/ liquid nitrogren, the sectioned in a cooled microtome
• used in biopsies

Question 12

Staining

Answer 12

• provides contrast so that the sample can be seen easier w/ the microscope
• also enhances certain features

Question 13

Histochemical staining

Answer 13

general staining, nonspecific

Question 14

Special staining

Answer 14

selectively stain certain cellular components of particular molecules like an enzyme
- important in diagnosis of some diseases

Question 15

Basophilic cell component staining

Answer 15

• negatively charged molecules
• methylene blue, toluidine blue, hematoxylin
• stains intense in the nucleus and some cytoplasmic granules

Question 16

Acidophilic cell component staining

Answer 16

• positively charged molecules
• eosin
• will stain more intensely in the cytoplasm, the plasma membrane, and some cytoplasmic membranes

Question 17

H&E Stain

Answer 17

• hematoxylin and eosin
• most common stain
• hematoxylin - stains dark blue/purple the nucleus, ribosomes, RER
• eosin - stains pink/red the cytoplasm, collagen

Question 18

Methylene blue

Answer 18

stains nucleus and ER/golgi apparatus blue

Question 19

Silver stain

Answer 19

stains certain proteins and delicate structures dark brown/black

Question 20

Osmium

Answer 20

stains lipids dark brown/black

Question 21

Periodic acid (Schiff)

Answer 21

stains complex carbs (basement membrane for ex.) red

Question 22

Light Microscopy

Answer 22

• specimen illuminated by visible light
• Resolving power = 0.2 micrometers
• Magnifies specimen up to 1000X

Question 23

Fluorescence microscopy

Answer 23

• specimen is stained w/ fluorescent dye, or a fluorescent-tagged antibody is bound to the specimen
• only detects one wavelength/color at a time
• Resolving power = 0.2 mm
• Mag up to 1000X

Question 24

Electric microscopy

Answer 24

• specimen stained w/ e- dense compound and lighten with e- beam
• magnets are used as lenses

Question 25

Transmission EM

Answer 25

• beam goes through the specimen
• ultrathin sections required (40-90 nm)
• resolving power 0.1-3 nm
• Mag up to 120,000X
• highest magnification of any type of microscopy, shows detailed structure of cellular organelles
• many organelles were discovered this way

Question 26

Scanning EM

Answer 26

• scanning the surface of the specimen, gives you a 3D view of the surface of the specimen
• specimen coated in Au or other e- dense ion
• does not require sectioning

Question 27

Immunocytochemistry

Answer 27

• specific type of histochemistry
• antibodies are used to detect specific proteins, and are tagged w/ fluorescent molecule
• antibodies binds to specific protein and binds to it when incubated w/ sample, making the protein complex visible using the tag on the antibody y

Question 28

Direct Immunocytochemistry

Answer 28

• antibody is tagged w/ chromophore of fluorescent molecules
• when the antibody binds to its target, it can be directly visualized (no extra steps required)

Question 29

Indirect immunocytochemistry

Answer 29

• primary antibody binds to target, but it doesn’t have a label, so you have to use a secondary, labeled antibody that detects the primary antibody

Question 30

How is Immunocytochemistry used in medical diagnosis?

Answer 30

• epithelial tumors
• breast cancer, colon cancer
• ductal cell tumor

Question 31

Artifact

Answer 31

• not present in normal tissue, interfere with interpretation of specimens
• EX: distortion of tissue during processing, shrinkage during fixation, separation of layers, folding/wrinkling, precipitation of stain, dust on slide