Immunohistochemistry (IHC | F)

Question 1

Provide a sample case (in relation to the application of IHC) and provide the clinical impression for the said case

Answer 1

A pt (male; 39 yo) has the ff:

1) Bilateral pleural effusion
2) Enlarged mediastinal lymph nodes
3) Enlarged paraaortic and retrocrural lymph nodes

Clinical impression: Bilateral Pleural Effusion secondary to Non-Hodgkin’s Lymphoma

Question 2

What are the immunostains that are suggested by the patho to be done in relation to their initial dx (w/ regards to the sample case) w/c is undifferentiated carcinoma vs. non-Hodgkin’s lymphoma?

Answer 2

Immunostain for:

1) Cytokeratin
2) Leukocyte common antigen (LCA)

Question 3

What are the 2 methods for immunohistochemical detection methods?

Answer 3

Can use:

1) Fluorescent probes
2) Enzymatic methods

Question 4

What is the principle of immunofluorescence technique?

Answer 4

This is where fluorescent substances (/ fluorophores) are used

Question 5

What is the principle of immunoenzyme technique?

Answer 5

Through enzymatic conversion (often by horseradish peroxidase [HRP]) of a soluble substrate (chromogen) into a non-soluble and colorful rxn product

Question 6

What are the steps for IHC (for enzymatic method)?

Answer 6

1) Tissue sections
2) Ag retrieval
3) Blocking endogenous enzymes
4) Secondary Ab
5) Primary Ab
6) Microscopy observation
7) Chromogen substrate
8) Counterstain
9) Mounting
10) Microscopy observation (by the patho)

Question 7

What are the applications of IHC in terms of lab dx of CA (immunocytochemistry)?

Answer 7

1) Application of immunochemical rxns on tissue sections / cell preparations
2) An ancillary technique employed in the dx of tumors
3) Demonstration of Ags (w/c are markers of certain tumors)

Question 8

What is the general principle of IHC?

Answer 8

Ag (protein trying to be detected) is present + primary Ab (attaches to the epitope) = Ag-Ab rxn

Question 9

What are the common applications of IHC?

Answer 9

1) Categorization of “undifferentiated” malignant tumors
2) Detection of Ags of potential prognostic / therapeutic importance
a. ER / PR (estrogen receptor / progesterone receptor)
b. Oncogene and tumor suppressor gene products
3) Determine the site of origin of metastatic tumors
4) Subclassification of tumors in various organ systems and tissue compartments
5) Categorization of leukemias and lymphomas
6) Distinction between adenocarcinoma (CK 8-18+) and mesothelioma (CK 8-18 -)

Question 10

What are the staining patterns in IHC?

Answer 10

1) AE1/AE3: shows (+) tumor cell staining
a. For small cell carcinoma
b. For IHC
2) Chromogranin: shows (+) cytoplasmic staining
a. For small cell carcinoma
b. For IHC
3) CD56: (+) w/ a membranous pattern
a. For small cell carcinoma
b. For IHC
4) TTF-1: shows diffuse (+) nuclear staining
a. For small cell carcinoma
b. For IHC
5) Ki-67 (panel d): shows a high proliferation rate w/ almost 100% tumor cell staining
a. For small cell carcinoma
b. For IHC

Question 11

What is the screening algorithm for the differentiation and categorization of malignant tumors?

Answer 11

1) The patho must 1st develop a differential dx
2) Differential dx depends on the tumor’s:
a. Histologic appearance
b. Anatomic location
c. Clinical setting

Question 12

What is primary screening panel and what is its use?

Answer 12

It is the “first-line” marker / Ab used to categorize undifferentiated malignant tumors accdg to histologic lineage

Question 13

What are the usual components of primary screening panel (for undifferentiated CAs)?

Answer 13

1) Pancytokeratin
2) LCA
3) S-100
4) Placental alkaline phosphatase (PLAP)
5) Synaptophysin
6) Vimentin

Question 14

Pancytokeratin is used for what?

Answer 14

Carcinoma

Question 15

LCA is used for what?

Answer 15

Lymphomas

Question 16

PLAP is used for what?

Answer 16

Germ cell tumor

Question 17

Synaptophysin is used for what?

Answer 17

Neuroendocrine (chromogranin)

Question 18

Vimentin is used for what?

Answer 18

Sarcoma (desmin and actin [can be added if pt has enough resources: for the differential dx to be more sure])

Question 19

What is the fxn of secondary screening panel?

Answer 19

To classify tumors accdg to histologic type

Question 20

Provide an ex. of the application of secondary screening panel (for lymphomas)

Answer 20

Pt is (+) for LCA -> it should be differentiated if it is CD 20 (+) / CD 3 (+) -> if it is CD 20 (+), it is a B cell type; if it is CD 3 (+), it is a T cell type

Question 21

Provide another ex. of the application of secondary screening panel (for anaplastic and super undifferentiated lymphomas)

Answer 21

Pt is (+) for LCA -> differentiate if pt is CD 20 (+) / CD 3 (+) -> if pt is CD 20 (+): it is B cell and/or lymphocyte predominant HL | if pt is CD 3 (+): it is T cell

But if pt is (-) for LCA -> the differential dx can be: 1) lymphoblastic lymphoma, 2) large cell anaplastic lymphoma, 3) immunoblastic lymphoma, 4) Hodgkin’s lymphoma (RS: CD15 [+]); except lymphocyte predominant (LCA [+])

Question 22

What is the relationship between pancytokeratin and LCA (in terms of dx)?

Answer 22

If pancytokeratin is (-) and LCA is (+): it is assured that it is lymphoma

But if both pancytokeratin and LCA are (-): tertiary screening panel is done

Pancytokeratin: for carcinoma
LCA: for lymphoma

Question 23

What is the differential workup if both cytokeratin and LCA are (-)?

Answer 23

Cytokeratin and LCA are both (-) -> can be 1) anaplastic lymphoma, 2) immunoblastic lymphoma, and/or 3) lymphoblastic lymphoma -> anaplastic lymphoma: CD 30 (+) | immunoblastic lymphoma: CD 20 (+) | lymphoblastic lymphoma: B-cell (CD 19 [+]) or T-cell (CD 3 [+])

Question 24

True or False

IHC is only an adjunct to dx

Answer 24

True

Question 25

True or False

Results (of / via IHC) must not be interpreted in the context of other findings (such as routine H & E sections and the clinical setting)

Answer 25

False, because results (of / via IHC) must be interpreted in the context of other findings (such as routine H & E sections and the clinical setting)