QUIZ Flashcards
T/F: The enzyme label for immunoperoxidase methods contains horseradish peroxidase
True
T/F: Monoclonal antibodies are often preferred over polyclonal antibodies because there is little bath-to-batch variability
True
T/F: DAB is an alcohol soluble chromagen
False
Synthetic resins may be used with DAB as it can be dehydrated and cleared.
What is the most common fixation/processing protocol for IHC?
Formalin fixed paraffin embedded (FFPE)
T/F: Harris hematoxylin cannot be used with AEC because of the alcohol present
True
Regulations regarding predictive marker staining of tissue places responsibility for the fixation time documentation on the lab
True
T/F: Precut control slides may be stored at room temperature indefinitely.
False
Must store in fridge
T/F: The procedure for staining cytology smears is the same as for routine slides.
False
Each slide must have its own protocol on the stainer
In the bloody areas of a tissue section stained with the immunoperoxidase technique, there is a marked reaction with RBCs. What happened?
Failure to use hydrogen peroxide
Paraffin sections stained with the immunoperoxidase technique show excessive background staining. What happened?
Nonimmune serum was not applied
If the first protein solution applied to the tissue is the primary antibody, nonspecific binding can occur
The skin control section for S100 was stained with the immunoalkaline phosphatase technique using fast red TR as the chromogen. It shows negative staining. What happened?
Sections were accidentally dehydrated and cleared causing the chromogen to breakdown
T/F: There is a negative correlation between extended cold ischemic time and loss of antigen ability.
False
The correlation is positive - longer times means worse staining
T/F: Prediluted antibody should always be used as provided by the manufacture
False
Always need to be validated before use
T/F: Negative tissue antigen controls may be run by substituting for the primary antibody the diluent in which the antibody is prepared
True
Negative controls omit the primary antibody
T/F: One standard protocol may be written to cover all specimens
False
Procedures need to be written for all non-standard tissue protcols
T/F: Zinc formalin preserves immunoreactivity very well
True
T/F: Multilink antibodies can only be used with monoclonal primary antibodies
False
Can be used with both mono and poly antibodies
T/F: Blocking reactions are used to block endogenous activity of the same enzyme used for the enzyme immune complex
True
Alpha, delta, gamma, and epsilon are all examples of:
Light polypeptide chains
Allergens
Epitopes
Heavy polypeptide chains
Heavy polypeptide chains
An antigen is a substance that triggers the production of:
immunogens
epitopes
hormones
immunoglobulins
immunoglobulins
Cells that engulf and destroy foreign substances and microorganisms are called:
Phagocytes
Myeloma Cells
Eosinophils
Natural Killer Cells
Phagocytes
A molecule which stimulates an immune response is called a(n):
immunoglobulin
epitope
immunogen
determinant
immunogen
Monoclonal antibodies are usually produced in which animal?
Cows
Goats
Mice
Cats
mice
Which of the following is a true statement about polyclonal antibodies?
Polyclonal antibodies are less sensitive than monoclonal antibodies.
Polyclonal antibodies produce less background staining than monoclonal antibodies.
Polyclonal antibodies can bind to multiple epitopes on one antigen.
Polyclonal antibodies are harvested from the spleen or lymph node of the host that they are produced in.
Polyclonal antibodies can bind to multiple epitopes on one antigen.
Antibodies are produced by:
B lymphocytes
Myeloma cells
Rabbit fusion partner cells
Phagocytes
B lymphocytes
Another term for background staining is:
enhanced staining
maximum staining
nonspecific staining
fluorescent staining
nonspecific staining
B cells are harvested with myeloma cells to produce:
hybirdomas
fusion cells
natural killer cells
rabbit fusion partner cells
hybirdomas
When using an IHC alkaline phosphatase staining method, what is the mechanism that produces the color?
An enzyme that hydrolyzes the substrate to phenolic compounds and phosphates. The phenols couple to colorless diazonium salts (chromogen) to produce insoluble, colored azo dyes.
An enzyme that catalyzes the reduction of hydrogen peroxide (H2O2) to water and oxygen.
An enzyme dissolves red food dye to tint the section
An enzyme that hydrolyzes the substrate to phenolic compounds and phosphates. The phenols couple to colorless diazonium salts (chromogen) to produce insoluble, colored azo dyes
A histotech is performing a CD-20 stain and receives a false-negative result on the specimen. What could be a cause for this happening?
Under fixation
T/F: Additives in alternative fixatives can cause either false-negative or false-positive immunohistochemical (IHC) staining.
True
Which of the following is an ADVANTAGE to using a polymer or micro polymer detection system?
Background free (typically) staining
Pretreatment of tisse for ISH can affect which two factors?
Signal strength
Background staining
T or F: The higher the temperature of the HIER solutions, the more effective the recovery of the epitope is
True
T/F: Most antigen retrieval methods apply temperatures near the freezing point of water.
False: boiling point
T/F: T or F: Certain non-epithelial tumors also stain positive for keratin
True
T/F: Vimentin is a sensitive marker for muscle differentiation and can be used to identify tumors derived from smooth, skeletal and cardiac muscle.
False: ACTIN
T/F: Melanomas and schwannomas always stain positive for vimentin
True
T/F: Vimentin has limited use as a stand-alone stain, but it can be very helpful when combined with other specific tumor markers.
True
T/F:T Neurofilament (NF) is most widely used to confirm the diagnosis of astrocytoma
False: Glial fibrillary acidic protein (GFAP)
Tumors that show neuronal or neuroendocrine differentiation will stain positive for what other marker?
neurofilament
A combination of keratin and chromogranin positivity is typical of _______
neuroendocrine carcinoma
Chromogranin positivity with a negative keratin stain is typical of ________
paraganglioma
T/F: PLAP is positive in majority of seminomas.
True
_______ are derived from neural crest and will be reactive for S100 protein
Melanocytes
The intensity of staining for S100 is usually ______________ proportional to the melanin content of the tumor.
Inversely
The best screening marker for lymphoma is _____.
LCA (leukocyte common antigen)
CD45
What are the immunophenotypic subclassification of lymphoma?
T cells (CD3, CD4, CD5)
B cells (CD19, CD20, CD23)
Reed-Sternberg cells (CD15, CD30)
immunoglobulin light and heavy chains
What is the most common immunohistochemical markers used to assess proliferation of tumor cells?
Ki-67 (MIB-1) and PCNA (proliferating cell nuclear antigen)
T/F: Chromogenic IHC staining can generate dense deposits that are easy to detect but difficult to quantitate.
True
T/F: Chromogenic detection is considered to be a less sensitive method than immunofluorescence.
False: more sensitive
The optimal incubation time for linking antibodies with peroxidase conjugates is ______________ at room temperature
30 to 60 minutes
T/F: Direct technique is less sensitive compared to indirect techniques.
True
T/F: The indirect technique of immunohistochemistry is a 2 or 3 step procedure that involves application of the unconjugated primary antibody, followed by a labeled antibody directed against the first antibody
True
In the indirect technique, cross-reactivity is eliminated by using _______.
preabsorbed secondary antiserum
What would be the purpose of the addition of a third layer of antibody?
To amplify the signal
T/F: Proteolytic enzyme digestion is necessary in renal and skin biopsies
True
Combining horseradish peroxidase with the most common chromogen, diaminobenzidine (DAB), results in a stable ________________ end product when antigen is present in the tissue.
insoluble dark brown reaction
In the PAP method, to block endogenous peroxidase activity, the sections are pre-incubated in ________ containing hydrogen peroxide
absolute methanol
Alkaline phosphatase antibodies raised in __________ , can also be used to form alkaline phosphatase-anti-alkaline phosphatase complexes (APAAP)
Mouse
T/F: The major advantage of APAAP procedure compared to the PAP technique is the lack of interference from exogenous peroxidase activity.
False: endogenous peroxidase activity
Endogenous alkaline phosphatase activity is usually blocked by adding ________ to the substrate solution
levamisole
T/F: Monoclonal antibodies, compared to polyclonal antibodies, depend more on environmental factors such as pH and solute for optimum performance.
True
What method uses avidin (derived from egg white) because of its marked affinity for biotin?
Avidin-Biotin Complex technique
Biotinylated label and avidin or streptavidin, and are added together _________ before use.
30 mins
T/F: Cryostat sections give much better antigen preservation than paraffin sections
True
T/F: Snap-frozen fresh tissue, without supporting media, can be stored at -10°C for long periods of time without appreciable antigen loss.
False: -70°C
Prolonged storage of OCT-embedded frozen tissue will cause gradual loss of cellular antigens
True
What fixative is generally used as fixative to preserve the antigen, to destroy harmful infective agents, and to allow a wide range of primary antibodies to be employed without destroying many of the epitopes.
Acetone
T/F: Formalin fixed paraffin-embedded material appears to be the best choice in a diagnostic setting of ISH.
True
T/F: In an aqueous environment, the thermodynamically stable conformation of a nucleic acid is that of a single-stranded helix.
False: double-stranded helix
The commonly used denaturants in the ISH method are:
Heat
Formamide
In a hybridization assay, the two sources are the ___________ and ____________.
target (sample) and the probe nucleic acids
The process of searching a sample for specific nucleic acid sequences is termed ________________.
hybridization reaction
T/F: RNA is much more unstable than DNA
True
SBB is positive in:
Acute Myeloid Leukemia (AML)
SBB staining is possibly a little more sensitive for the early myeloid cells
True
T/F: Metamyelocytes stains negative for SBB
False: metamyelocytes stain strongle positive
A piece of skin has been stained according to a standard melanoma protocol. You notice that after counterstaining the visible IHC product has been removed. What most likely occurred to cause this result?
Incorrect pre-treatment used
Dehydrated in alcohol
Incorrect antibody dilution
Chromogen was outdated
Dehydrated in alcohol
In immunohistochemical staining of formalin-fixed tissue, proteolytic enzyme pretreatment of a tissue section:
enhances background staining
unmasks antigen
epitopesis needed to demonstrate all antigens
has precise end-points
unmasks antigen epitopes
Which of the following is NOT a section in the MSDS?
-PPE
-physical hazards
-primary routes of entry
-staining procedures
staining procedures
How many mls of a 1:5 dilution of 100% alcohol would be water?
180 ml
190 ml
200ml
160 ml
160 ml
160 ml
The red chromagen used in this image is particularly helpful for this type of specimen because it:
-Aids in diagnosis of melanocytic lesions
-Reduces background staining
-Is better for quantitative image analysis
-Is more sensitve
Aids in diagnosis of melanocytic lesions
What is the staining pattern in the image?
-Nuclear
-Nuclear and cytoplasmic
-Membranous
-Cytoplasmic
Nuclear and cytoplasmic
This is an image of Melan-A staining using alkaline phosphatase detection with a red chromagen. The tissue is skin. Which of the following statements is true?
The tissue is stained appropriately. Melan-A has nuclear visualization.
The tissue is stained inappropriately. Melan-A has neither cytoplasmic visualization or nuclear visualization
The tissue is stained inappropriately. Melan-A has neither cytoplasmic visualization or nuclear visualization
The tissue is stained inappropriately. Melan-A has neither cytoplasmic visualization or nuclear visualization
This image is of prostate tissue stained with P63 antibody. Which of the following statements is true?
The tissue is stained inappropriately. P63 is a membranous marker, but there is also aberrant nuclear visualization.
The tissue is stained inappropriately. The basal cells are not staining with P63.
The tissue is stained inappropriately. P63 stains basal cells with nuclear visualization, but there is also aberrant membranous staining.
The tissue is stained appropriately.
The tissue is stained inappropriately. P63 stains basal cells with nuclear visualization, but there is also aberrant membranous staining.
Which of the following immunostains would be appropriate to request on this tissue section?
Cytokeratin 7, cytokeratin 20, and TTF-1
High molecular weight keratin and PSA
CD 3, CD5, CD20, and CD79a
Estrogen receptor, progesterone receptor and HER2/neu
Cytokeratin 7, cytokeratin 20, and TTF-1
The staining present in this specimen is the result of:
-lymphocytes stained with leukocyte common antigen (LCA)
- inadequately blocked biotin in red blood cells
-inadequately blocked horseradish peroxidase in red blood cells
-inadequately blocked biotin in lymphocytes
inadequately blocked horseradish peroxidase in red blood cells
Identify the chromogen used in the image which is insoluble in alcohol and other organic substances.
-diaminobenzidine
-3amino - 9 - ethylcarbazole
-4 chloro 11 napthol
-diaminopropylene
diaminobenzidine
The red chromagen used in this image is particularly helpful for this type of specimen because it:
-Aids in diagnosis of melanocytic lesions
-Reduces background staining
-Is better for quantitative image analysis
-Is more sensitive
Aids in diagnosis of melanocytic lesions
