ASCP Immunohistochemistry 2 — Questions and Answers
Question 1: What does a negative IHC result on a poorly fixed specimen most likely represent?
- True absence of the antigen
- False negative due to antigen destruction by inadequate fixation (Correct answer)
- Successful antibody staining with no target present
- Over-retrieval artifact
Correct answer: False negative due to antigen destruction by inadequate fixation
Inadequate fixation causes protein degradation that destroys antigenic epitopes, resulting in false-negative IHC results even when the antigen is present.
Question 2: In IHC, what is the function of the blocking step with normal serum?
- To fix the tissue before antibody application
- To prevent non-specific binding of the secondary antibody to tissue proteins (Correct answer)
- To enhance primary antibody penetration
- To amplify signal detection
Correct answer: To prevent non-specific binding of the secondary antibody to tissue proteins
Normal serum (from the species used to raise the secondary antibody) saturates non-specific protein binding sites, reducing background staining.
Question 3: Ki-67 IHC is used to assess what cellular property?
- Apoptosis rate
- Cellular proliferation index (Correct answer)
- Hormone receptor status
- Lymphoid lineage
Correct answer: Cellular proliferation index
Ki-67 is a nuclear protein expressed only in proliferating cells; IHC for Ki-67 measures the proportion of actively dividing cells (proliferation index).
Question 4: What is the difference between the direct and indirect IHC methods?
- Direct uses enzyme-labeled primary antibody; indirect uses unlabeled primary and enzyme-labeled secondary antibody (Correct answer)
- Direct uses two antibodies; indirect uses one
- Direct is more sensitive than indirect
- There is no practical difference
Correct answer: Direct uses enzyme-labeled primary antibody; indirect uses unlabeled primary and enzyme-labeled secondary antibody
In the direct method, the primary antibody carries the detection label; in the indirect method, an unlabeled primary antibody is detected by a labeled secondary antibody for greater sensitivity.
Question 5: What IHC marker is used to confirm the presence of muscle differentiation in a tumor?
- CK7
- Desmin or smooth muscle actin (SMA) (Correct answer)
- CD20
- TTF-1
Correct answer: Desmin or smooth muscle actin (SMA)
Desmin and smooth muscle actin are cytoskeletal proteins expressed in muscle cells; their IHC positivity confirms muscle differentiation in tumor cells.
Question 6: Over-retrieval in heat-induced antigen retrieval (HIER) can cause which artifact?
- Absence of all staining
- Section detachment from the slide (Correct answer)
- Increased specific staining only
- Blue background staining
Correct answer: Section detachment from the slide
Excessive heat or prolonged retrieval times can cause sections to detach from slides, especially if adhesion is suboptimal.
ASCP Immunohistochemistry 2
What does a negative IHC result on a poorly fixed specimen most likely represent?