Histology Wk 1

Question 1

What is histology

Answer 1

The study of tissues of the body

Question 2

What are the two interacting components of tissues

Answer 2

Cells and extra cellular matrix (ecm)

Question 3

What does the ecm consist of…

Answer 3

Many kind of macromolecules which form complex structures such as collagen fibrils

It supports cells

Question 4

Name the steps for preparation of tissue for study

Answer 4

Fixation
Dehydration
Clearing
infiltration
Embedding
Trimming

Question 5

What is fixation

Answer 5

Small pieces of tissue are placed in solutions of chemicals that cross link proteins and inactivate degradative enzymes, which preserve the tissue structure.

Because fixative must fully diffuse through the cell it is cut into small fragments before fixation to facilitate penetration

Question 6

What is dehydration

Answer 6

The tissue is transferred through a series of increasingly concentrated alcohol solutions ending in 100 percent which removes all water

Question 7

What is clearing

Answer 7

Alcohol is removed in organic solvents in which both alcohol and paraffin are miscible

Question 8

What is infiltration

Answer 8

The tissue is then placed into melted paraffin in an oven at 52-60 degrees until it becomes completely infiltrated with this substance

Question 9

What is embedding

Answer 9

The paraffin infiltrated tissue is placed in a small mold with melted paraffin and allowed to harden

Tissues are embedded with plastic resin are also dehydrates in ethanol and then dehydrated in ethanol and infiltrated with plastic solvents that harden when cross linking polymerisers are added

Question 10

What is trimming

Answer 10

The resulting paraffin block is trimmed to expose the tissue for sectioning on a microtome

Question 11

What is a microtome

Answer 11

It is used for sectioning paraffin embedded tissues for light microscopy

Question 12

What are paraffin section usually cut into

Answer 12

3-10 micrometers thickness for light microscopy and less than 1micrometer for electron microscopy

Question 13

What is the term basophilic

Answer 13

Cell components such as nucleus acids with negative negative charge have an affinity for basic dyes

Question 14

What is the term acidophillic

Answer 14

Cationic components such as proteins with many ionized amino groups stain more readily with acidic dyes

Question 15

Name some acidophilic and basophilic components

Answer 15

Basophilic
DNA
Rna
Glycoaminoglycans

Acidophilic
Mitochondria
Secretory granules
Collagen

Question 16

What is hematoxylin used for

Answer 16

Stains dna in the cell nucleus, rna rich portions of the cytoplasm and the matrix of the cartilage producing a dark blue or purple color

Question 17

What is eosin used for

Answer 17

Stains cytoplasmic structures and collagen pink

Question 18

Describe the period acid- Schaffer reaction (PAS)

Answer 18

Utilises the hexose rings of polysaccharides and other carbohydrate rich tissue structures and stains such macromolecules purple or magenta

Question 19

What is autoradiography

Answer 19

It is a method of localizing newly synthesized macromolecules in cells or tissue sections

Question 20

What are radioactively labeled metabolites

Answer 20

Nucleotides
Amino acids
Sugars

Question 21

How are slides with radioactively labeled metabolites produced

Answer 21

Slides containing these are coated in a darkroom with photographic emulsion in which silver bromide crystals act as micro detectors of the radiation

Silver bromide crystals are reduced by radiation to produce small black grains of metallic silver either under LM OR TEM which indicate the locate the radio labelled macromolecules in the tissue

Question 22

What is in vitro in cell and tissue culture

Answer 22

Live cells can be maintained and studied outside the body in culture

Question 23

What is in vivo in cell and tissue culture

Answer 23

Cells are bathed in fluid derived from blood plasma and containing many different molecules required for survival and growth.

Question 24

What is enzyme histochemistry

Answer 24

Is a method for localizing cellular structures using specific enzymatic activity present in those structures

Question 25

What are the steps for enzyme histochemistry

Answer 25

1. Tissue sections are immersed in a solution containing the substrate of the enzyme to be localized 2. The enzyme is allowed to act on its substrate 3. The section is then put on contact with a marker compound that reacts with a product of the enzymatic action on the substrate 4. The insoluble structure is visible by light or electron miscroscopy

Question 26

Visualizing specific molecules

Answer 26

Labels are fluorescent compounds, radioactive atoms that can be detected with autoradiography Molecules of peroxidase or other enzymes can be detected with histochemistry Metal particles can be seen with light and electron microscopy

Question 27

What is immunohistochemsitry

Answer 27

A highly specific interaction between macromolecules is that between an antigen and its antibody To localize and identify many specific proteins

Question 28

What is an advantage of using monoclonal antibodies rather than polyclonal antibodies

Answer 28

It can be selected to be highly specific and bind strongly to the protein to be detected

Question 29

What is the method of immunohistochemistry

Answer 29

A tissue section that one believes contains the protein of interest is incubated in a solution containing antibody against the protein The antibody binds specifically to the protein and after a rinse the protein’s location in the tissue can be seen with a light of electron microscope LOOK AT NOTEBOOK IN GOODNOTES

Question 30

What are the two methods of immunocytochemistry

Answer 30

Direct and indirect

Question 31

What is the direct method

Answer 31

If the cell or tissue antigen of interest is detected by directly binding a labeled primary antibody specific for that antigen Fluorescent antibodies bind to the direct target antibody

Question 32

What is the indirect method

Answer 32

Uses an unlabelled primary antibody that is detected bound to its antigen with labeled secondary antibodies

Question 33

Why is the indirect method used more widely in research and pathologic tests

Answer 33

It is more sensitive with the extra level of antibody binding serving to amplify the visible signal and cheaper

Question 34

What is hybridisation

Answer 34

The specific binding between two single strands of nucleus acid, which occurs under appropriate conditions if the strands are complementary

Question 35

What is in situ hybridisation

Answer 35

When nucleic acid sequences in solution are applied directly to prepared cells and tissue sections

Question 36

What is in situ hybridisation ideal for

Answer 36

Determining if a cell has a specific sequence of dna such as a gene Identifying the cells containing specific mRNA Determining the localisation of a gene in a specific chromosome

Question 37

Probes

Answer 37

The nucleotide sequence of interest is detected with probes The probe is obtained by cloning by polymerase chain reaction

Question 38

Why is Sudan black used

Answer 38

Lipid soluble

Question 39

What fixative is used in light microscopy

Answer 39

37% formaldehyde

Question 40

What fixative is used in EM

Answer 40

Glutaraldehyde reacts with the amine groups of proteins