IHC Simplified Final Review

Question 1

Self vs Non-self

Answer 1

in immune response

Question 2

What do Immunizations do?

Answer 2

Prevent disease

Question 3

Pasteurization

Answer 3

kills bacteria, tested attenuated anthrax and cholera in chickens to demonstrate the concept of acquired immunity and immunization

Question 4

Antigens

Answer 4

“foreign invaders” that trigger an immune response

Viruses, bacteria, foreign skin grafts, pollen

Question 5

Innate vs Adaptive immunity

Answer 5

Innate is general

Adaptive has “memory” and is more specific, develops through li fee experience/exposure

Question 6

T cells

Answer 6

Attack infected cells, viruses, and bacteria

Mature in the thymus

Question 7

B cells

Answer 7

Secrete antibodies that attach to antigens

Work against most bacteria

Mature in the bone marrow

Question 8

Epitope specificity

Answer 8

Epitope of the antigen vs paratope of the antibody

Question 9

Antibody structure

Answer 9

2 heavy chains (constant region)

2 light chains (specific region) paratope

Question 10

Monoclonal ABs

Answer 10

Hybridomas made from aggressive myelomas and a single antibody clone of desirable affinity from a mouse injected with the target antigen

Question 11

Advantages of monoclonal ABs

Answer 11

More specific, less background, no lot to lot or batch to batch variation

Question 12

2 types of AB light chains

Answer 12

kappa or lambda

Question 13

3 stages of tumor malignancy

Answer 13

Induction
In situ
Invasion

Question 14

Meaning of immunohistochemistry

Answer 14

Immuno: antigen/antibody
Histo: tissue based
Chemistry: reaction

Question 15

all the basic steps for an indirect immuno

Answer 15

Fixation, embedding, sectioning
Antigen retrieval
Endogenous enzyme blocking
Background/non-specific stain blocking
Primary AB
Secondary AB
Chromogen
Counterstain
Dehydrate, mount, coverslip
Read the slide

Question 16

Two types of antigen retrieval

Answer 16

Heat and enzyme

Question 17

Enzymes for retrieval

Answer 17

proteinase K, trypsin, pepsin

Question 18

Hydrogen peroxide is used to block endogenous enzyme activity

Answer 18

in horseradish peroxidase protocols

Question 19

Horse radish peroxidase is an enzyme that

Answer 19

has a substrate: hydrogen peroxide

Question 20

Review how to do dilution math

Answer 20

.

Question 21

Cold ischemic time

Answer 21

time between removal from the body and submersion in fixative, ideally 1 hour (or less if possible)

Question 22

Fixation times for HER2 and ER

Answer 22

6 hours to 72 hours to not mess up IHC staining

Question 23

How to deparaffinize

Answer 23

Xylene->100%EtOH->95%EtOH->H2O

Question 24

Semi-quantitative scoring on PD-L1

Answer 24

PD-L1
0, 1+, 2+, 3+, 4+ based on percent of positive cells within a field of view
Only membrane staining is positive, both partial and complete count as positive
Count only cancer cells, not inflammatory cells because macrophages are frequently positive for PD-L1
Tumor cells are bigger than normal macrophages
Any intensity is positive
Intensity can vary within the tumor
Reporting % positive cells is important and you must have at least 100 viable tumor cells for the count to be considered valid

Question 25

What a pathology report should include

Answer 25

all patient demographics (name, DOB, MR#)
Tissue identifiers
ICD10 code for billing
All IHC stain results both positive and negative
Differential diagnosis to justify stains ordered
Frozen vs paraffin origin/fixation/processing
Antibodies used
Interpretation of results

Question 26

What is ISH (in-situ hybridization)

Answer 26

Detection of mRNa in cells/tissue by hybridizing a single stranded DNA or RNA probe via complimentary base pairing to a target sequence in tissue.