QUIZ Flashcards by Mary Williams

T/F: The enzyme label for immunoperoxidase methods contains horseradish peroxidase

True

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T/F: Monoclonal antibodies are often preferred over polyclonal antibodies because there is little bath-to-batch variability

True

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T/F: DAB is an alcohol soluble chromagen

False

Synthetic resins may be used with DAB as it can be dehydrated and cleared.

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What is the most common fixation/processing protocol for IHC?

Formalin fixed paraffin embedded (FFPE)

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T/F: Harris hematoxylin cannot be used with AEC because of the alcohol present

True

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Regulations regarding predictive marker staining of tissue places responsibility for the fixation time documentation on the lab

True

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T/F: Precut control slides may be stored at room temperature indefinitely.

False

Must store in fridge

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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

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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

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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

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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

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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

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T/F: Prediluted antibody should always be used as provided by the manufacture

False

Always need to be validated before use

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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

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T/F: One standard protocol may be written to cover all specimens

False

Procedures need to be written for all non-standard tissue protcols

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T/F: Zinc formalin preserves immunoreactivity very well

True

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T/F: Multilink antibodies can only be used with monoclonal primary antibodies

False

Can be used with both mono and poly antibodies

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T/F: Blocking reactions are used to block endogenous activity of the same enzyme used for the enzyme immune complex

True

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Alpha, delta, gamma, and epsilon are all examples of:

Light polypeptide chains

Allergens

Epitopes

Heavy polypeptide chains

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An antigen is a substance that triggers the production of:

immunogens

epitopes

hormones

immunoglobulins

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Cells that engulf and destroy foreign substances and microorganisms are called:

Phagocytes

Myeloma Cells

Eosinophils

Natural Killer Cells

Phagocytes

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A molecule which stimulates an immune response is called a(n):

immunoglobulin

epitope

immunogen

determinant

immunogen

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Monoclonal antibodies are usually produced in which animal?

Cows

Goats

Mice

Cats

mice

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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.

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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

After an alkaline phosphatase immunostain run was completed, the slides were manually counterstained in hematoxylin, dehydrated, cleared and coverslipped. The entire run was negative. What is the most likely source of error? -the wrong type of hematoxylin was used as counterstain -the primary antibodies were all omitted -the counterstaining solution masked alkaline phosphatase activity -the end-product for alkaline phosphatase activity was dissolved by alcohol

the end-product for alkaline phosphatase activity was dissolved by alcohol

Of the following, the preferred fixative for demonstration of intermediate filaments contains: alcohol formalin mercury dichromate

alcohol

According to OSHA, which of the following records can be kept for less than 30 years? exposure records for specified carcinogens fit testing results for personal chemical respirators medical records for exposure to hazardous chemicals personal protective equipment training

fit testing results for personal chemical respirators

A biotin block is often used as part of an IHC staining protocol for the reason of suppressing staining of endogenous biotin. All of the following are tissues that require biotin blocking except: Kidney Liver Thyroid Brain

Thyroid

When staining for neuroendocrine tumors the most beneficial markers of identification are: s-100 and cytokeratin chromogranin and synaptophysin neurofilament and neuron specific enolase neuron specific enolase

chromogranin and synaptophysin

The primary advantage for using non-formalin fixatives in immunochemistry it to: reduce exposure to toxic chemicals prevent loss of antigenicity better preserve tissue morphology reduce cost

prevent loss of antigenicity

Which statement is true concerning pH? a liquid is more acidic if its pH is higher adding sodium hydroxide will make the pH higher indicator paper always turns blue if the pH is high the pH increases by 2 units for a 10?C temperature rise

adding sodium hydroxide will make the pH higher

mmunologic stains and flow cytometry are performed on malignant cells found in pleural effusion specimen. The following results are obtained: B72.3 positive; carcinoembryonic antigen (CEA) positive; DNA content aneuploid. Based on this profile, the cells most likely represent: reactive mesothelial cells malignant mesothelioma metastatic adenocarcinoma Hodgkin's lymphoma

metastatic adenocarcinoma

A section labeled with an antibody has no background but the antigen of interest stains very weakly. This is an example of: low specificity, high sensitivity high specificity, low sensitivity high specificity, high sensitivity low specificity, low sensitivity

high specificity, low sensitivity

As a chromogenic substrate, amino-ethylcarbazole (AEC) would be preferred to diaminobenzidine (DAB): if permanent preparation is desired when staining pigmented lesions to reduce contact with a potential carcinogen for subsequent metallic color enhancement

when staining pigmented lesions

A common fluorochrome used in immunohistochemistry is: fluorinated isotryptophan fluorine isothiocyanate fluorescein isothiocyanate fluoroisothiocyanate

fluorescein isothiocyanate

The chemical used to quench endogenous alkaline phosphatase in tissue sections is: biotin hydrogen peroxide levamisole EDTA

levamisole

The regulatory agency responsible for establishing and enforcing laws concerning the disposal of chemicals down the drain is: EPA GLP NIOSH NTP

EPA

The primary base units used in the metric system are: liters, grams, meters ounces, grams, meters pounds, gallons, meters meters, grams, pints

liters, grams, meters

A single plasma cell (activated B-cell) produces: all the antibodies necessary to neutralize foreign antigens polyclonal antibodies against a specific antigen antibodies specific to a single antigenic determin antantibodies specific to hypervariable determinants

antibodies specific to a single antigenic determinant

A negative control consisting of normal animal serum shows background staining. The primary slide, however, shows strong specific staining with no background. The likely cause of this discrepancy is that the normal serum: was more dilute has antibodies to something in the tissue, while the primary serum does not was not an absorbed antiserum, while primary is an absorbed antiserum had exceeded the expiration date, and probably contained a contaminant microorganism

has antibodies to something in the tissue, while the primary serum does not

Convert 3.5 gallons to liters 12.5 liters 14 liters 13.247 liters 13 liters

13.247 liters

Fluorochromes are available in several colors. In order to see these colors, a fluorescence microscope must be equipped with: a color video camera brightfield illumination the correct wavelength filters a quartz halogen bulb

the correct wavelength filters

In the indirect immunohistochemistry staining method, the: test slide is first treated with unlabeled antiserum test slide is first treated with enzyme-labeled to the primary antibody second step is treatment with enzyme labeled antiserum to the primary antigen second step is treatment with unlabeled antiserum to the primary antibody

second step is treatment with enzyme labeled antiserum to the primary antigen

When selecting reagents for peroxidase-anti-peroxidase (PAP) staining, the PAP complex should be prepared in the same (or closely related) animal species as the: chromogen secondary antibody primary antibody bridging antibody

primary antibody

The specific substrate of horseradish peroxidase is: diaminobenzidine ethylene diamine tetra acetic acid hydrogen peroxide polyethylene glycol

hydrogen peroxide

B-lymphocytes can produce specific antibodies without T-lymphocyte help only if the antigen is: processed by macrophages a cluster designation greater than 10,000 kd T-independent

T-independent

The primary reason that frozen sections destined for immunohistochemistry are fixed in some way after sectioning is to: keep the tissue from falling off the slide thaw the tissue in a controlled manner prevent diffusion of the tissue antigens of interest kill any infectious organisms that may present

prevent diffusion of the tissue antigens of interest

The best positive control tissue for IHC is one that is: bought from a reputable vendor as fresh as possible processed identically to patient tissue is available in large quantities

processed identically to patient tissue

When staining sections for examination by immunofluorescence microscopy: tissue antigens are "stained" by fluorescent-labeled antibodies tissue antibodies are "stained" by fluorescent-labeled antigens fluorescent dyes are sandwiched between antigen and antibody fluorescent dyes are bound to tissue with heavy metal mordants

tissue antigens are "stained" by fluorescent-labeled antibodies

The ability of an antibody to recognize and bind with a specific protein antigen is due to its: molecular size carbohydrate structure the dimensional structure animal species of origin

the dimensional structure

After fixation for 24 to 48 hours in formalin, tissue for IHC should be stored in which of the following fluids to preserve antigenicity? 10% buffered formalin 50% acetone 70% ethanol Hanks buffered saline

70% ethanol

Which of the following are enzymes commonly used in immunohistochemistry to unmask antigenic determinants? papain, propane, chymase, lactase pepsin, tryptophan, kinase, proteinase K pronase, trypsin, proteinase K, pepsin lactase, pepsin, trypsin, pepsin

pronase, trypsin, proteinase K, pepsin

Which of the following is a chromogen used in peroxidase histochemistry that generates a product that is soluble in alcohol or xylene but not in water? 9-aminoethylcarbazole (AEC) 3,3'-diaminobenzidine (DAB) alpha-naphthyl phosphate (ANP) hexazonium pararosaniline (HPR)

9-aminoethylcarbazole (AEC)

During QC of an IHC stain it is noticed that a particular marker has not stained and needs to be repeated. The block, however, has been depleted. All of the following are acceptable ways to re-stain the specimen except: Staining an unstained slide with a positive control Staining over a negative control Staining a previous HE of the same specimen Staining over the IHC slide that did not work

Staining over the IHC slide that did not work

Formalin fixation causes cross-linkages, what affect might this have on IHC staining? Masks epitopes enhances staining allows antigens to bind faster and easier removes the need for pre-treatment

Masks epitopes

Staining of a blood or marrow smear for peroxidase :ctivity is sometimes helpful in the diagnosis of myeloid leukemia, but intensity of staining is sometimes low. To ensure that the incubation has been adequate: incubate a control slide that has been smeared with a material known to have enzyme activity, such as a peroxidase-labeled antiserum incubate until some staining is seen, because there is always some stainable peroxidase activity, even in normal leukocytes treat the smear with 1% hydrogen peroxide in methanol for 10 minutes before incubating in the peroxidase reaction medium incubate a control smear with added sodium azide to inhibit peroxidase

incubate until some staining is seen, because there is always some stainable peroxidase activity, even in normal leukocytes

he 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 cellsin adequately blocked biotin in lymphcytes

inadequately blocked horseradish peroxidase in red blood cells

In immunoperoxidase staining, efficiency of the chromogen-substrate reaction step can be checked by: doubling the methanol, hydrogen peroxide concentration extending the time in the substrate-chromogen increasing the concentration of chromogen omitting the blockage of endogenous peroxidase

omitting the blockage of endogenous peroxidase

Antibodies must always be stored at: 2-8 C 2-6C 1-8 C according to manufacturers specifications

according to manufacturers specifications

Commercial monoclonal antibodies are produced by all of the following except: synthesized utilizing automated instrumentation in a factory a single plasma cell and its clones mice in a lab cell culture

synthesized utilizing automated instrumentation in a factory

Heat induced epitope retrieval (HEIR) may be used for: all tissue sections, for any antibody all formalin-fixed tissue sections those antibodies for which it has been determined to be optimal frozen sections

those antibodies for which it has been determined to be optimal

With regard to the avidin-biotin- complex (ABC) immunohistochemistry method: the primary antiserum is avidin-labeled no conjugation steps are involved the ABC complex binds to biotin-labeled secondary antibody bridging antiserum is added in excess

the ABC complex binds to biotin-labeled secondary antibody

The specific substrate of horseradish peroxidase or HRP is: diaminobenzidineethylene diamine tetraacetic acid hydrogen peroxide polyethylene glycol

hydrogen peroxide

A chemical group which gives the property of color to a chromogen is a: chromatophore chromatography chromophore chromogranin

chromophore

Monoclonal antibodies are highly specific but may show low sensitivity because: the dilution is too high the specific binding site in the tissue may be altered of the difficulty of preparing monoclonals of poor technique by the technologist

the specific binding site in the tissue may be altered

Optimization can best defined as The clinical process of confirming that the 'reactivity' that is observed The technical process of adjusting various conditions in order to produce the greatest amount of specific staining (sometimes referred to as 'signal'). Developing an IHC staining protocol that definitively confirms that the results produced are 'biologically 'correct' The process of documenting a protocol

The technical process of adjusting various conditions in order to produce the greatest amount of specific staining (sometimes referred to as 'signal').

Antibody diversity is due to different amino acid sequences of the: variable regions of the heavy chain variable regions of the light chain variable regions of both heavy and light chains Fc regions of the heavy chains

variable regions of both heavy and light chains

Which of the following is the substrate of the enzyme peroxidase? HRP DAB H2O2 TRIS

H2O2

In peroxidase staining, the colored end product is formed following: reduction of a chromogenic substrate and oxidation of horseradish peroxidase reduction of hydrogen peroxide and oxidation of a chromogenic substrate oxidation of hydrogen peroxide and reduction of a chromogenic substrate oxidation of horseradish peroxidase and reduction of hydrogen peroxide

reduction of hydrogen peroxide and oxidation of a chromogenic substrate

Which of the following is true of a polyclonal antibody? lot to lot consistency sustained production reacts with a specific epitope reacts with multiple epitopes on a single molecule

reacts with multiple epitopes on a single molecule

Because of the harshness of IHC procedures on tissue slides, which of these factors in slide preparation are very important to a high quality IHC slide? Charged slides Drying time of tissue on slides Well fixed tissue All of the above

All of the above

Which one of the following is most correct regarding heat induced epitope retrieval (HIER) techniques? Steamers, pressure cookers, microwaves and waterbaths can all be successfully used for HIER. Lead thiocyanate formic acid and water have all been successfully used as HIER solutions. You cannot "over retieve" tissue using HIER techniques. Both "Steamers, pressure cookers, microwaves and waterbaths can all be successfully used for HIER." and "Lead thiocyanate formic acid and water have all been successfully used as HIER solutions." are correct.

Both "Steamers, pressure cookers, microwaves and waterbaths can all be successfully used for HIER." and "Lead thiocyanate formic acid and water have all been successfully used as HIER solutions." are correct.

In immunohistochemical procedures using horseradish peroxidase (HRP), a solution of hydrogen peroxide is used to: block endogenous peroxidase enhance background staining affect reactivity of antibodies and antigens intensify the coloring reaction

block endogenous peroxidase

An appropriate immunohistochemical reaction is dependent upon: incubation time incubation temperature antibody dilution All of the above

All of the above

A section of intestine is labeled with an anti-keratin antibody and visualized with alkaline phosphatase. The section shows strong staining along the entire brush border of the intestinal epithelium. What artifact must you be aware of while interpreting this stain? nothing, the intestine shows background staining with all chromogens anti-keratin antibody often stains collagen and smooth muscle the intestinal brush border cells contain endogenous alkaline phosphatase formalin pigment can mimic the pattern of keratin staining in the intestine

the intestinal brush border cells contain endogenous alkaline phosphatase

An anaplastic metastatic malignant tumor is submitted for diagnosis with the request to rule out malignant melanoma. The best antibody panel to study this problem should include: vimentin, cytokeratin, S-100 and HMB45 cytokeratin, S-100, CEA, EMA HMB-45, actin, desmin, CD99 S-100, chromogranin, vimentin, NSE

vimentin, cytokeratin, S-100 and HMB45

An immunohistochemistry labeling method in which a molecule of avidin is directly labeled with an enzyme is called: avidin-biotin complex labeled avidin-biotin peroxidase-antiperoxidase alkaline phosphatase-antialkaline phoshatase

labeled avidin-biotin

Formalin fixation causes cross-linkages, what affect might this have on IHC staining? Masks epitopes enhances staining allows antigens to bind faster and easie removes the need for pre-treatment

Masks epitopes

Which of the following panel results are improbable? Keratin +, vimentin -, LCA -, S100- Keratin +, Vimentin +, LCA +, S100 + Keratin -, Vimentin +, LCA -, S100 + Keratin -, Vimentin +, LCA - S100 -

Keratin +, Vimentin +, LCA +, S100 +

n IHC, a negative control is used to: determine what steps contribute to any non-specific staining show what the tissue would stain like if it was negative test the dilution of the primary antibody save primary antibody

determine what steps contribute to any non-specific staining

Following immunoperoxidase staining, it is observed that the positive control stains well, but there is no staining of a specimen expected to be positive. Of the following, the most likely cause is that: the specimen was improperly counterstained the specimen was not properly fixed staining steps were not performed in the correct order excess buffer or nonimmune serum diluted the primary antibody

the specimen was not properly fixed

Which statement about the pH of a solution is true? the pH number is the concentration of hydrogen ions in gram-ions or moles per liter a concentrated acid can have a negative pH, such as -1.5 the pH is a meaningful value for all liquids, even those that contain no water a solid object, such as a crystal, a hair or a section of tissue has a pH value

a concentrated acid can have a negative pH, such as -1.5

In immunohistochemical staining of formalin-fixed tissue, proteolytic enzyme pretreatment of a tissue section: enhances background staining unmasks antigen epitopes is needed to demonstrate all antigens has precise end-points

unmasks antigen epitopes

A "checkerboard" titration of primary vs. secondary antibodies is used to determine the: best primary antibody concentration only best secondary primary antibody concentration optimal combination of concentrations of each antibody optimal chromogen concentration for each antibody

optimal combination of concentrations of each antibody

A single plasma cell (activated B-cell) produces: all the antibodies necessary to neutralize foreign antigens polyclonal antibodies against a specific antigen antibodies specific to a single antigenic determinant antibodies specific to hypervariable determinant

antibodies specific to a single antigenic determinant

Which of the following combinations would yield the best double-stain? DAB as chromogen with ER and new fuchsin as the chromogen with PR DAB as chromogen with kappa and new fuchsin as chromogen with lambda Fast red as chromogen with high molecular weight keratin and new fuchsin as chromogen with pan keratin Fast Blue as chromogen for CD3 and BCIP-NBT as chromogen for CD20

DAB as chromogen with kappa and new fuchsin as chromogen with lambda

Fixatives that tend to mask antigenic sites and hamper immunohistochemical localization of antigens contain: mercury phosphates aldehydes alcohol

aldehydes

The peroxidase staining procedure in which reagents are linked exclusively by antigen-antibody reactions without any conjugation steps is: avidin-biotin-peroxidase complex method peroxidase-anti-peroxidase indirect immunoperoxidase method direct immunoperoxidase method

peroxidase-anti-peroxidase

A brain biopsy reveals a poorly differentiated malignant tumor. The differential diagnosis is primary versus metastatic brain cancer. The most useful antibody panel to resolve this differential is: cytokeratin, vimentin, muscle common actin S-100, cytokeratin AE1/3, synaptophysin glial fibrillary acid protein, cytokeratin AE1/3, synaptophysin carcinoembryonic antigen, leukocyte common antigen, S-100

glial fibrillary acid protein, cytokeratin AE1/3, synaptophysin

If your antibody is FDA approved but your protocol is different how many tests do you need to run when validating? 10 previously known positive cases 20 positive and 20 negatives cases 10 negative cases 15 positive and 15 negative cases are run

20 positive and 20 negatives cases

The step in the IHC stainings process that allows the exposure of the sought after epitopes is called: direct staining detection antigen retrieval antibody matching

antigen retrieval

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

Paraffin sections for IHC should be cut at: 1 to 2 um 3 to 5 um 6 to 10 um 10 to 20 um

3 to 5 um

In order to assess the effects of over-fixation and the potential need for antigen retrieval on a given specimen, the inclusion of an antibody against what ubiquitous intermediate filament is suggested in all initial antibody panels? ubiquitin cytokeratin vimentin actin

vimentin

What marker is considered almost a universal control for over-fixation? keratin CD30 vimentin kappa

vimentin

The size of a tissue sample is important for immunohistochemistry because: smaller samples are easier to section the fixative must penetrate the entire sample larger samples are easier to handle larger samples will stay on the slide better

the fixative must penetrate the entire sample

DAB may be enhanced by the use of: Copper, silver, gold or iodine Cobalt, nickel, silver or gold Potassium, sodium, silver or gold Cobalt, iodine, copper or nickel

Cobalt, nickel, silver or gold

In immuno-alkaline phosphatase staining, which control is useful to determine the level of endogenous alkaline phosphatase? incubate in chromogen only incubate overnight in levamisole apply all steps except the enzyme stain several slides at different time periods

incubate in chromogen only

Fluorochrome-conjugated primary antibodies are an example of: direct labeling indirect labeling the ABC system the PAP system

direct labeling

the specific binding site in the tissue may be altered

the ABC complex binds to biotin-labeled secondary antibody

mmunoperoxidase-stained tissues show reaction of red blood cells and granulocytes with the chromogenic substrate. The most likely explanation is that: they contain antigen that reacted with the primary antiserum the specimens were improperly counterstain edendogenous peroxidase was not blocked specimens were allowed to dry during the staining procedure

endogenous peroxidase was not blocked

The primary advantage for using non-formalin fixatives in immunochemistry it to: reduce exposure to toxic chemicals prevent loss of antigenicity better preserve tissue morphology reduce cost

prevent loss of antigenicity

omitting the blockage of endogenous peroxidase

The three critical parameters of the heat-mediated antigen retrieval procedure are: correct microwave wattage, pH, and volume proper temperature, incubation time, and pH volume of solution, temperature, incubation time incubation time, pH, and style of steamer

proper temperature, incubation time, and pH

A metastatic carcinoma of unknown origin is presented for diagnosis. All of the following are useful to determine the primary site of the tumor except: vimentin cytokeratin 7 and 20 CEA S-100

S-100

All enzymes: change the rate of chemical reactions catalyze oxidation-reduction reactions act upon extracellular substrates have enhanced activity at 60C

change the rate of chemical reactions

The two principle defense mechanisms of the body's immune system are: humoral and enzymatic hydrolytic and cell-mediated humoral and cell-mediated hydrolytic and enzymatic

humoral and cell-mediated

Following avidin-biotin-peroxidase complex staining, the cytoplasm of scattered single cells in the stroma of both positive and negative control slides is stained. The most likely explanation is that the: cells represent avidin-containing mast cells primary antibody was applied to all slides endogenous peroxidase was not blocked staining steps were not performed in the correct order

cells represent avidin-containing mast cells

has antibodies to something in the tissue, while the primary serum does not

when staining pigmented lesions

peroxidase-anti-peroxidase

synthesized utilizing automated instrumentation in a factory

To eliminate the problem of endogenous staining a particular IHC method is optimal. What method would that be? LSAB+ ABC Polymer-Based PAP

Polymer-Based

A cryostat may be defined as a/an: device that maintains a controlled low temperature instrument that cuts sections of frozen specimens microtome contained in a freezing cabinet section cut from a frozen specimen

microtome contained in a freezing cabinet

Light chain monoclonality is the hallmark of: B-cell non-Hodgkin's lymphoma Hodgkin's disease Hairy cell leukemia T-cell non-Hodgkin's lymphoma

B-cell non-Hodgkin's lymphoma

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

The antibody class most frequently demonstrated in immunofluorescent and immunoenzyme staining is: IgA IgE IgG IgM

IgG

A cell antigen that can be demonstrated by immunofluorescent staining of fresh-frozen tissues is not visible by immunoperoxidase staining of formalin-fixed paraffin-processed tissue. The difference in the procedures that most likely accounts for the lack of staining is: masking of antigen binding sites by fixation difference in chromogen the sequence of staining steps interpretation under different microscopy systems

masking of antigen binding sites by fixation

those antibodies for which it has been determined to be optimal

The specific substrate of horseradish peroxidase or HRP is: diaminobenzidine ethylene diamine tetraacetic acid hydrogen peroxide polyethylene glycol

hydrogen peroxide

In immunohistochemical staining, a limitation of polyclonal antisera as opposed to monoclonal antisera is: that the production of polyclonal antibodies is technically more difficult the greater cross-reactivity of polyclonals with similar antigens the limited availability of polyclonal antisera the extreme specificity of polyclonal antibodies

the greater cross-reactivity of polyclonals with similar antigens

Polymer based detection uses: A structure made of polymer and biotin that contains secondary antibodies A backbone of polymer with the addition of enzymes A polymer backbone with secondary antibodies and hydrogen peroxide incorporated in the structure A polymer backbone that includes secondary antibodies and enzymes

A polymer backbone that includes secondary antibodies and enzymes

Which of the following is a very useful positive control for lymphatic IHC stains? stomach tonsil kidney brain

tonsil

Sensitized T-cells respond to antigenic challenge by secreting a series of biological mediators termed: prostaglandins cytokines kinins c-reactive proteins

cytokines

test slide is first treated with unlabeled antiserum

S-100 protein shows reactivity with all of the following except: Schwann cells and glial tissue chrondrocytes and adipocytes smooth muscle and skeletal muscle Langerhans' histiocytes and interdigitating reticulum cells

smooth muscle and skeletal muscle

Individuals who are hypersensitive to environmental allergens usually have above normal circulating levels of: IgE IgG IgM IgD

IgE

When using an alkaline phosphatase detection system: It is always necessary to perform a hydrogen peroxidase quench It is never necessary to perform a hydrogen peroxidase quench, but you must always perform an alkaline phosphatase quench. It is recommended that an alkaline phosphatase quench be performed on frozen tissues Using a quench of any type will result in very weak staining when using an alkaline phosphatase detection system

It is recommended that an alkaline phosphatase quench be performed on frozen tissues

All of the following are intermediate filaments except: Vimentin Keratin Glial fibrillary protein Actin

Actin

To eliminate the problem of endogenous staining a particular IHC method is optimal. What method would that be? LSAB+ ABC Polymer-Based PAP

Polymer-Based

adding sodium hydroxide will make the pH higher

9-aminoethylcarbazole (AEC)

masking of antigen binding sites by fixation

Name two preservatives used in commercial preparation that can reduce antibody reactivity

Sodium chloride Azides

What buffer should not be used as an antibody diligent unless specifically recommended by the manufacturer?

PBS

Archival unstained sections are stained for ER and PR because the original slides were lost; the newly stained slides were negative for both ER and PR, despite having been reported previously as positive. What is the likely cause of the discrepancy? tissues were overfixed in formalin improper antigen retrieval method was used the sections are too old the ER and PR antibodies have expired

the sections are too old

Polyclonal antibody serum often labels non-specific antigens in the tissue because polyclonal: -antibodies can shift their hypervariable regions to conform to other epitopes -antibodies are stickier than monoclonal antibodies -antiserum is thicker than monoclonal serum -antiserum contains many antibodies

antiserum contains many antibodies

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

Aids in diagnosis of melanocytic lesions