Immunohistochemistry Flashcards
An antigen is a substance that triggers the production of:
immunoglobins
A light chain present in some antibodies is:
kappa
Different molecular sites on antigens are known as:
epitopes
When an antibody labeled with a chromogen is reacted with tissue from a patient, the IHC technique is called:
direct
Which of the following is a fluorochrome:
a. alkaline phosphatase
b. rhodamine
c. fast red
d. diamineobenzidine
rhodamine
In the indirect IHC method the…
patients serum is added to tissue sections containing known antigens
In the avidin-biotin methods, the primary antibody is followed by:
biotinylated secondary antibody
A dye that absorbs light and then emits its own light at a longer wavelength is known as:
a fluorochrome
In some IHC tehcniques, alkaline phosphatase functions as the:
enzyme
Horseradish peroxidase is used in some avidin-biotin methods as the:
enzyme
A substrate for alkaline phosphatase-labeled antibodies is commonly:
napthol-AS-phosphate
Which of the following chromogens is INSOLUBLE in alcohol:
a. fast red TR
b. 3,3’-diaminobenzidine tetrahydrochloride (DAB)
c. 3-amino-9-ethylcarbazole (AEC)
d. fast blue BBN
b. 3,3’-diaminobenzidine tetrahydrochloride (DAB)
IF 3-amino-9-ethylcarbazole is used as a chromagen, the hematoxylin used for counterstaining should be:
alcohol free
Tissue for immunoflourescence must be:
frozen and unfixed
A neoplasm is defined as a/an:
new growth of uncontrolled cell multiplication
What is NOT a proper approach to validation of a new antibody?
using only the manufacturers recommendations
When the vimentin stain is completely negative on formalin-fixed tissues, this indicates that most likely the tissue has been:
overfixed
Besides the heat employed, another important factor in head-induced epitope retrieval (HIER) us the:
composition of the solution used
Ficin is used in IHC for:
enzyme-induced epitope enhancement
What has been eliminated with polymeric IHC staining methods?
serum and avidin-biotin blocking steps
In the peroxidase-antiperoxidase (PAP) method of antigen detection, the PAP complex is made in the same species as:
the primary antibody
Imidazole may be used in peroxidase techniques to:
intensify the DAB reaction
What link antibodies would most likely follow as a monoclonal primary antibody?
rabbit anti-mouse
T/F: Prediluted antibodies should always be used as provided by the mansufacturer.
False.
T/F: Negative tissue antigen controls may be run by substituting for the primary antibody the diluent in which the antibody is prepared.
True
T/F: One standard staining protocol may be written to cover all specimens.
False.
T/F: Zinc formalin preserves immunoreactivity well.
True.
T/F: Multilink antibodies can be used only with monoclonal primary antibodies.
False.
T/F: Blocking reactions are used to block endogenous activity of the same enzyme as that used for the enzyme immune complex.
True
T/F: The enzyme label for immunoperoxidase methods contain horseradish peroxidase.
True
T/F: Monoclonal antibodies are often preferred over polyclonal antibodies because there is no batch-to-batch variability.
True.
T/F: DAB is an alcohol-soluble chromagen.
False.
T/F: Harris hematoxylin should not be used with 3-amino-9-ethylcarbazole (AEC) because of the alcohol present.
True.
T/F: Regulation regarding predictive marker staining of the tissue places responsibility for documenting time in the fixative on the laboratory.
True.
T/F: The only method of heat-induced epitope retrieval involves using a microwave oven.
False.
T/F: Optimal dilutions for each antibody must be determined by your lab.
true
T/F: Metal salts involving nickel, copper, and osmium may be used to intensify 3-amino-9-ethylcarbazole reaction.
False. Metal salts intensift DAB.
T/F: Alkaline phosphatase label can be substituted for peroxidase label in most IHC methods.
True
T/F: Precut control slides may be stored at room temp indefinitly.
False.
In the bloody areas of a tissue section stain with the immunoperoxidase technique, there is a marked reaction of the red blood cells. This is most likely the result of:
failure to use hydrogen peroxide. It is essential to reduce non-specific staining of RBCs.
Paraffin sections stained with the immunoperoxidase technique show excess background staining. What could explain this?
Forgetting to apply the non-immune serum. If the first protein solution applied to the tissue is the primary antibody, nonspecific binding can occur.
The skin control for S-100 was stained with the immunoalkaline phosphatase technique using fast red TR as the chromogen. It shows negative staining. One possible explanation is:
sections were dehydrated and cleared. With prolonged dehydration, the alkaline phosphatase chromogen can break down.
At the end of the immunoperoxidase staining procedure, it was realized that an anti-goat linking antibody was used with monoclonal HMB-45 antibody. Microscopic results should show a:
negativity stained positive control and a negative stained specimen since monoclonal antibodies are most often prepared by mice or rabbits. An anti-mouse, anti-rabbit or multilink linking secondary must be used.
Both the known positive control and the specimen are negative following immunoperoxidase staining with LCA primary antibody. DAB was used as a chromogen. What could be the cause?
wrong secondary antibody was used. The secondary antibody must be targeted to the primary for linking to occur.
Excessive background is noted on both the control and specimen stained with immunoperoxidase. This might be due to:
insufficient washing with buffer
Excessive background is noted on the specimen but not the control stained with immunoperoxidase. This is most likely due to:
free antigen in the tissue
Both the positive control and the specimen stained with immunoalkaline phosphatase using CEA antibody show very weak staining. Why?
incorrectly diluted antibody.
