Fixation Flashcards
An example of an additive fixative is one that contains:
a) Picric acid
b) Acetic acid
c) Ethyl alcohol
d) acetone
A
When compared with tissue fixed in formalin, tissue fixed in zinc-formalin will show:
a) better ultrastructural preservation
b) decreased immunoreactivity
c) increased enzyme activity
d) superior nuclear detail
D
Zinc is not as toxic as mercury and has been substituted for mercury-containing fixatives, as well as added to formaldehyde, because of the comparable, or increased, nuclear detail and increased antigenicity.
Microscopic examination of an H&E stained section fixed in formalin shows marked nuclear bubbling. One most often sees this artifact if the specimen is processed following:
a) incomplete fixation
b) prolonged fixation
c) microwave fixation
d) frozen sectioning
A
If tissue is incompletely fixed before placing in the dehydrating solution, a nuclear bubbling artifact may result.
Microscopic evaluation of H&E stained sections from a surgically removed small bowel specimen shows an absence of much of the epithelium in otherwise normal tissue. This most likely resulted from:
a) mechanical trauma
b) delayed fixation
c) ulceration
d) poor choice of fixative
B
When prolonged delay in fixation occurs, some cells may completely disappear, such as the epithelial cells in the intestinal tract. GI specimens should be opened, pinned out, and placed in fixative immediately upon receipt.
A specimen of kidney must be shipped to another city for immunofluorescence studies. The specimen should be placed in:
a) saline
b) Michel solution
c) buffered formalin
d) Orth solution
B
Tissue for immunofluorescence studies must be unfixed and when unfixed tissue is to be held for several days or transported over a long distance, then Michel transport medium is recommended.
A certain project requires a fixative that contains acetic acid yet stabilizes erythrocyte membranes. One fixative that could be used is:
a) Zenker solution
b) Bouin solution
c) Gendre solution
d) Hollande solution
D
The cupric acetate present in Hollande solution stabilizes RBC membranes, so that the lysis that occurs with Bouin solution, and other acetic acid containing fixatives, is much less.
When the microwave oven is used for fixation, the most critical factor is the:
a) preparation of the formalin solution
b) use of glass containers
c) control of the temperature
d) osmolality of the fixation solution
C
When the microwave is used for fixation, irreversible morphologic damage will result if the temperature is not carefully controlled.
To adequately remove the calcium from a specimen containing areas of microcalcification, the tissue could be fixed in:
a) Hollande solution
b) neutral buffered formalin
c) B-5 solution
d) Zamboni solution
A
Hollande solution is a modification of Bouin solution, which contains acetic acid, and thus will decalcify small specimens of bone. None of the other fixatives listed are acidic.
Which of the following fixatives contains copper acetate?
a)Hollande
b) Bouin
c) Gendre
d) Zamboni
A
Hollande solution contains cupric acetate
A specimen is submitted with the statement that was fixed in formalin. Microscopic sections show marked autolysis of erythrocytes. This indicates that the fixative most likely was:
a) prepared with too much formalin
b) buffered above neutrality
c) acidified with acetic acid
d) not formalin
C
Marked lysis of erythrocytes is characteristic of fixatives containing acetic acid.
Fixatives are classified as additive because of the:
a) addition of several chemicals to the solution
b) addition, or binding of the fixative to tissue proteins
c) additional reactions occurring with longer fixation
d) additional reactive tissue sites available for dye binding
B
Additive fixatives chemically link, or add themselves onto, the tissue and change it with this action.
Kidney biopsy tissue has been fixed in Phosphate-buffered glutaraldehyde for 2 hours and then placed in phosphate buffer solution. If a portion of this tissue is processed for light microscopy, sections would most likely show:
a) very poor glomerular preservation
b) decreased uptake of hematoxylin
c) lysis of cytoplasmic elements
d) nonspecific PAS staining
D
Glutaraldehyde is a dialdehyde, and the extra aldehyde group is not involved in most crosslinking reactions (fixation); therefore it is left free to react in any method using Schiff reagent, such as PAS stain - leading to false positives.
Uric acid crystals are preserved ONLY when tissue is fixed in:
a) absolute alcohol
b) neutral buffered formalin
c) Orth solution
d) Zamboni solution
A
Uric acid crystals are water soluble, so can be maintained in the tissue only with a fixative solution containing no water. Absolute alcohol is recommended.
Improper preservation of tissue will result if there is:
a) a delay in fixation
b) rapid penetration of the fixing fluid
c) prolonged storage following formalin fixation
d) rapid dehydration, clearing, embedding, and sectioning
A
A delay in fixation will cause improper preservation of the tissue. The nuclei may show a loss or complete disappearance of chromatin; the tissue may also show disruption of the cytoplasm, cell shrinkage, artifactual spaces around cells, or complete loss of some cells.
A good fixative will:
a) render cell constituents soluble
b) minimize differences in tissue refractive indices
c) protect tissue against alteration during subsequent processing
d) minimally affect tissue metabollic processes
C
A fixative should stabilize the tissue elements, so that the effect of any subsequent procedures, such as processing, will be minimal.
The function of methanol in commercial formalin solutions is to:
a) retard the polymerization of formaldehyde
b) prevent the formation of formic acid
c) stabilize the formalin at a basic pH
d) permit room temperature storage of formalin
A
Commercial formaldehyde contains about 10-14% methanol which is added to help prevent polymerization to paraformaldehyde, a highly polymeric form of formaldehyde.
The problem shown in this image is the result of: (see page 4 of study guide)
a) incomplete fixation
b) excessive dehydration
c) incomplete clearing
d) poor paraffin infiltration
A
The cracks in the tissue and the smudgy nuclei are due to incomplete fixation.
In electron microscopy, Zamboni fluid, gluteraldehyde, and osmium tetroxide function as:
a) dehydrating agents
b) clearing agents
c) embedding media
d) fixative solutions
D
Zamboni fluid, glutaraldehyde, and osmium tetroxide function as fixatives for specimens for electron microscopy. Osmium tetroxide may be used either as a primary or secondary fixative. Zamboni solution and glutaraldehyde are usually followed by osmium tetroxide to increase fat retention, and thus membrane preservation.
Tissue will remain unfixed if placed in:
a) potassium dichromate
b) sodium borate
c) osmium tetroxide
d) zinc chloride
B
Sodium borate is not a fixative.
Bouin solution is contraindicated (not good) for:
a) small tissue biopsies
b) tissue intended for subsequent trichrome stains
c) tissue to be stained by the Feulgen reaction
d) routine tissue sections
C
Picric acid is a sufficiently strong acid to hydrolyze nuclei, so if stains for DNA (Feulgen) or RNA are anticipated, any fixative containing picric acid (Bouin, Gendre, Hollande) should be avoided.
Formalin pigment can be removed from tissue sections by treatment with 10%:
a) hydrochloric acid in 70% alcohol
b) nitric acid in 70% alcohol
c) sulfuric acid in 70% alcohol
d) ammonium hydroxide in 70% alcohol
D
Formalin pigment can be removed by treating tissue sections with alcoholic picric acid or alkaline alcohol.
Stock neutralized formalin is prepared in the lab by storing the solution over a layer of calcium carbonate. The solution withdrawn from this stock container will:
a) become acidic
b) become alkaline
c) remain neutral
d) exhibit metachromasia
A
Solutions of neutralized formalin prepared by storing the solution over a layer of calcium carbonate will gradually become acidic, because the pH has not been stabilized by buffering the solution.
Microscopic evaluation reveals a poorly stained H&E section of spleen. These results will be difficult to remedy if the problem is:
a) poor fixation
c) improper sectioning
c) poor staining
d) incorrect section placement
A
When poorly fixed tissue is processed and embedded, the staining results are most often not optimum and are very difficult, if not impossible, to remedy.
To make 10% formalin solution, how many mL of water should be added to 300mL of 37%-40% formaldehyde solution?
a) 1,800
b) 2,500
c) 2,700
d) 3,600
C
300mL of 37-40% formaldehyde solution added to 2,700mL of water will yield a total of 3,000mL of a 10% formalin solution. Preparation includes diluting 1 part stock solution with 9 parts water.
One action of acetic acid is to:
a) exert a shrinking effect on tissue
b) render nucleoprotein acidophilic
c) form salt linkages between protein chains
d) coagulate tissue lipids
D
The major use of acetic acid in fixatives is the precipitation (coagulation) and preservation of nucleoproteins.
Aldehyde fixatives are used for electron microscopy preparations because they:
a) are readily available
b) visibly stain tissue
c) preserve cell ultrastructure
d) coagulate tissue lipids
C
Aldehyde fixatives are used for electron microscopy preparations because they preserve cell ultrastructure. The must be followed by secondary osmium tetroxide fixation to preserve lipids.
A fixative containing potassium dichromate:
a) is suitable when histochemical techniques are planned
b) will result in excellent subsequent silver staining
c) is preferred for the preservation of argentaffin cells
d) will make tissue more receptive to eosin staining
D
If mercuric chloride is used alone for fixation, it will:
a) leave tissue proteins uncoagulated
b) procedure a very acidic solution
c) penetrate poorly and cause excessive shrinkage
d) decrease tissue affinity for stains
C
Poor penetration and excessive shrinkage will result if mercuric chloride is used alone for fixation. It is a powerful protein coagulant and enhances staining by leaving the tissues very receptive to dyes.
Tissue stored for long periods of time in unbuffered formalin or in acetate formalin may show brown, crystalline pigment in stained sections. To remove this pigment prior to staining it is necessary to treat the section with:
a) saturated alcoholic picric acid
b) alcoholic lithium chloride
c) iodine & sodium thiosulfate
d) potassium permanganate & oxalic acid
A
The pigment is black acid hematin, or formalin pigment, which tends to form when the pH of the solution drops below 6.0; this may happen in unbuffered formalin solutions. The pigment may be removed by treating with saturated alcoholic picric acid.
For good fixation of tissue with osmium tetroxide for electron microscopy, it is recommended that the tissue segment be no lorger than:
a) 1mm^3
b) 2mm^3
c) 1cm^3
d) 2cm^3
A
Osmium tetroxide penetrates very poorly, and so specimens should be minced to approximately 1mm cubes for electron microscopy and cut very thin for demonstrating fat in paraffin sections.
The tissue shown in this image is: (see sage 6 of study guide)
a) liver
b) kidney
c) spleen
d) lymph node
C
Following fixation with Bouin solution, tissue should be washed with:
a) absolute alcohol
b) 50-70% alcohol
c) 20-40% alcohol
d) saline solution
B
Traditionally, tissue fixed in Bouin solution is washed in 50-70%, or 70% alcohol saturates with lithium carbonate, before processing. If excess picric acid is left in embedded tissue, the staining will deteriorate.
The PTAH staining technique would require postfixation, or mordanting, if the tissue were originally fixed in:
a) Bouin
b) Zenker
c) Gendre
d) formalin
D
PTAH stains are not good after formalin fixation. Zenker fixative has been used traditionally, but Bouin solution and other mordants are also effective.
The problem seen in this image is known as: (see page 7 of study guide)
a) cell shrinkage
b) smudgy nuclei
c) pyknotic nuclei
d) nuclear bubbling
D
Absolute ethanol is poor choice for the fixation of:
a) glycogens
b) pigments
c) lipids
d) blood smears
C
Absolute ethanol will dissolve lipids, and therefore should not be used if lipid preservation is important.
Which of the following fixatives may give false positive results in some carbohydrate techniques?
a) neutral buffered formalin
b) Bouin solution
c) Gendre solution
d) glutaraldehyde
D
Glutaraldehyde is a dialdehyde. The extra aldehyde body does not form crosslinks with the tissue, leaving one aldehyde group free to react in techniques depending on the demonstration of aldehydes (PAS).
It is necessary to adjust the pH of the most formalin solutions because of the presence of:
a) methanol
b) formic acid
c) paraformaldehyde
d) carbon dioxide
B
Formaldehyde solutions become acidic by reacting with atmospheric oxygen to form formic acid; therefore, most formalin solutions require raising the pH, or preferably buffering to approximate neutrality. Formic acid is undesirable because it leads to the formation of formalin pigment.
The rate of fixation varies with the fixative and also with the:
a) time placed in the fixative of choice
b) expected completion time of the report
c) anticipated special stains needed
d) temperature of the fixative solution
D
Carnoy solution is recommended for the preservation of:
a) acid-fast bacilli
b) nucleic acids
c) lipids
d) red blood cells
B
Carnoy solution exhibits good nuclear preservation but lyses red blood cells, dissolves lipids, and is not recommended for the preservation and subsequent demonstration of acid-fast bacilli.
Which of the following factors affects fixation for light microscopy the least?
a) temperature
b) volume ratio
c) penetration rate
d) pH
D
Good preservation of tissue for light microscopy is least dependent on fixative pH, and many fixatives are quite acidic. Varying the pH from 4 to 9 apparently makes little difference in the fine structure produced by formalin fixation; however, a pigment is produced at the lower pH. The other factors listed are very important.
Formalin pigment is generally created in tissue fixed in formalin when the pH:
a) rises above 6
b) falls below 6
c) is buffered to neutrality
d) is 7.2
B
Very bloody cytology smears are often treated with:
a) 10% formalin
b) Hollande solution
c) Clark solution
d) acetone
C
Because blood sometimes obscures important cellular detail in very bloody cytology smears, Clark solution is used to lyse the RBCs.
Glyoxal is one of the newer fixatives, which has the added advantage of:
a) ability to crosslink
b) rapidity of action
c) enhanced staining
d) preservation of erythrocytes
B
Glyoxal fixatives are extremely rapid in action, and surgical specimens are fixed after only 4-6 hours of exposure. Glyoxal forms crosslinks only under very specific conditions, lyses erythrocytes, and there may be a slight reduction in staining especially after long periods of storage
If a tissue was fixed in a solution different from that required for a staining procedure, microscopic sections frequently can be stained anyway if they are:
a) soaked in a solution of lithium carbonate prior to staining
b) revitalized by washing in a solution of sodium bisulfite
c) postfixed in the appropriate fixative prior to staining
d) treated with hydrogen peroxide
C
Sections can usually be mordanted, or postfixed, in the fixative required for yielding the best staining results
The nuclear problem seen in this image is: (see page 9 of study guide)
a) understained chromatin
b) cell shrinkage
c) smudgy nuclei
d) nuclear bubbling
C
Fixation in Bouin solution is:
a) recommended for the Feulgen reaction
b) excellent for ultrastructural preservation
c) the cause of considerable swelling of tissue
d) frequently used for endocrine tissues
D
Bouin solution is an excellent fixative for biopsy specimens of the gastrointestinal tract and of the endocrine system.
B-5 fixative contains:
a) mercuric chloride, sodium acetate, and glacial acetic acid
b) mercuric chloride, potassium dichromate, and glacial acetic acid
c) mercuric chloride, sodium acetate, and 37-40% formaldehyde
d) mercuric chloride, potassium dochromate, and 37-40% formaldehyde
C
Pigment caused by mercury-containing fixatives can be removed from tissues by:
a) saturated alcoholic picric acid
b) iodine-sodium thiosulfate
c) washing in running water
d) potassium hydroxide in water
B
Iodine followed by sodium thiosulfate is used for the removal of pigment caused by mercury.
Which of the following fixatives has a mordanting effect on tissue?
a) Carnoy solution
b) 10% calcium formalin
c) absolute alcohol
d) Bouin solution
D
Bouin solution is indicated as a mordant for trichrome stains if the tissue was not originally fixed in that solution.
Tissue should be placed in a fixative solution immediately after removal from the body to:
a) prevent decomposition due to enymatic activity
b) permit the dehydrant to function preoperly
c) inhibit crosslinking of the tissue proteins
d) stabalize tissue carbohydrates
A
Decomposition of tissue by enzymes begins as soon as the blood supply is interrupted; therefore the tissue should be placed in fixative immediately after removal.
Calcium-formalin fixative is recommened for the BEST preservation and subsequent demonstration of:
a) glycogen
b) phospholipids
c) amyloid
d) estrogen receptors
B
For most fixatives, the volume of fixing fluid in relation to the volume of tissue should be:
a) 2-5 times
b) 6-9 times
c) 10-14 times
d) 15-20 times
D
Ultrastructural preservation will be very poor following fixation in:
a) Zamboni PAF
b) 2% buffered gluteraldehyde
c) osmium tetroxide
d) 10% aqueous formalin
D
Glyoxal is a/an:
a) aliphatic hydrocarbon
b) aromatic hydrocarbon
c) dialdehyde
d) ketone
C
Zinc-formalin fixatives:
a) give poor ultrastructural preservation
b) can be used to preserve enzymes
c) result in poor nuclear detail
d) will not coagulate tissue proteins
A
Zamboni PAF refers to a fixative containing:
a) potassium dichromate, acetic acid, & formaldehyde
b) potassium aluminum sulfate & paraformaldehyde
c) buffered picric acid & formaldehyde
d) picric acid, acetic acid, & formaldehyde
C
The problem seen in this image most likely could have been prevented by: (see page 11 of study guide)
a) immediate & prolonged fixation
b) less processing time
c) increased paraffin infiltration
d) better microtomy technique
A
The preferred fixative when tissue is to be stained for the presence of simple fats is:
a) Zenker
b) Helly
c) Hollande
d) neutral buffered formalin
D
Fixation of cytology smears should occur within:
a) 1-2 seconds
b) 10-15 seconds
c) 40-45 seconds
d) 1 minute
A
When osmium tetroxide is used as a fixative in histology, it:
a) destroys lipids
b) interfered with staining
c) leaves tissue very soft
d) distorts cell membranes
B
The breakdown of tissue due to enzyme activity is called:
a) polymerization
b) putrefaction
c) autolysis
d) osmosis
C
The fixative of choice for the demonstration of a gouty tophus is:
a) neutral buffered formalin
b) absolute alcohol
c) Bouin solution
d) Zenker solution
B
A good fixative for routine use is one that:
a) makes tissue more penetrable to fluids
b) is hypotonic to the tissue constituents
c) enhances putrefaction of tissue components
d) promotes tissue autolysis
A
A pigment caused by chromate-containing fixatives can be prevented by treating the tissue prior to processing with:
a) running water
b) iodine
c) picrid acid
d) potassium permanganate
A
When fixing tissue for electron microscopy with formaldahyde or gluteraldehyde, the preservation of ultrastructure depends upon all of the following EXCEPT the:
a) pH
b) time and temperature
c) concentration and purity of the reagent
d) type of tissue
D
Formic acid present in commercial formalin solutions may:
a) facilitate pigment formation
b) precipitate hemosiderin
c) promote staining
d) cause tissue shrinkage
A
Carnoy solution is a combination of which of the following chemicals:
a) absolute alcohol, acetone, & glacial acetic acid
b) cedarwood oil, absolute alcohol, & glacial acetic acid
c) acetone, chloroform, & absolute alcohol
d) chloroform, glacial acetic acid, & absolute alcohol
D
When using a nonimmunologic stain for chromaffin granules, it is necessary to fix the tissue in a:
a) mercury fixative
b) primary chromate fixative
c) formalin fixative
d) picric acid fixative
B
When liver tissue is fixed with 2-3% gluteraldehyde:
a) glycogen is dissolved
b) penetration rate is very rapid
c) chemical reaction occurs with lipids
d) the ultrastructure is preserved
D
A poor fixative is characterized by:
a) the absence of shrinkage or swelling of tissue
b) inactivation of tissue enzymes
c) slow tissue penetration
d) the absence of distortion or dissolution
C
The problem seen in this image possible could have been prevented by: (see page 13 of study guide)
a) buffering the formaldehyde solution
b) immeiate contact with fixation solution
c) grossing the specimen more carefully
d) better processing protocol
B
Bouin solution contains all of the following EXCEPT:
a) picric acid
b) absolute alcohol
c) 37-40% formaldehyde
d) glacial acetic acid
B
Coagulant fixatives:
a) change the spongework of proteins into a mesh-like network
b) produce fewer artifacts than noncoagulant fixatives
c) act very slowly to fix tissue
d) leave protein linkages unaffected
A
The breakdown of tissue by bacterial action is called:
a) autolysis
b) putrefaction
c) act slowly to fix tissues
d) leave protein linkages unaffected
B
When ultrastructural preservation is of the utmost importance, the fixative used should have a pH of:
a) 6.8 - 7.1
b) 7.2 - 7.4
c) 7.6 - 7.8
d) 8.0 - 8.2
B
A fixative component that produces a diffuse brownish black pigment is:
a) picric acid
b) osmium tetroxide
c) mercuric chloride
d) acetic acid
C
For the BEST preservation of staining properties during long-term storage, tissues should be stored in:
a) buffered formalin
b) 10% formal-saline
c) 70% ethanol
d) Zamboni solution
C
Ethanol is useful as a fixative because it:
a) crosslinks proteins
b) increases tissue basophilia
c) prevents tissue shrinkage
d) preserves glycogen very well
D
The nuclear problem seen in this image is the most often attributed to: (see page 14 in study guide)
a) delay in fixation
b) incomplete fixation
c) overdehydration
d) poor paraffin infiltration
B
To prevent the formation of formalin pigment in tissues, formalin should be:
a) heated
b) cooled
c) buffered
d) acidified
C
Which of the following fixatives should be used for specimens that may NOT be processed for several days?
a) 10% neutral buffered formalin
b) Bouin solution
c) Helly solution
d) Zenker solution
A
A biopsy that was placed in water by mistake is submitted to the lab. This mistake most likely will cause:
a) mushy sections
b) swollen and ruptured cells
c) hardening of the tissue
d) no appreciable changes
B
Sections of breast CA were fixed in a saline solution in the microwave oven. Microscopic examination of H&E show marked pyknotic, overstained nuclei. The staining results were most likely caused by the:
a) use of saline for fixation
b) solution temp exceeding 68C
c) use of plastic containers in microwave
d) presence of CA in the breast tissue
B
An unknown pigment in a tissue section that can be bleached with a saturated alcoholic solution of picric acid is most likely:
a) melanin pigment
b) formalin pigment
c) hemosiderin
d) mercury pigment
B
The formaldehyde in Helly solution:
a) causes reduction of some chemicals in the solution
b) coagulates and denatures tissue proteins
c) prevents turbidity and precipitate formation
d) eliminates the need for postfixation washing
A
Formaldyhyde solutions for routine use are most commonly buffered by:
a) monobasic & dibasic phosphates
b) sodium acetate & acetic acid
c) s-collidine & hydrochloric acid
d) sodium barbitol & sodium hydroxide
A
The preferred fixatie for the image seen below is: (see pg 16)
a) 10% neutral buffered formalin
b) alcohol
c) Clark solution
d) Bouin solution
D
One characteristic of Zamboni fixative is that it:
a) does not stabilize cellular proteins
b) may be used for electron microscopy
c) is easily destroyed by tissue fluids
d) must be followed by osmium tetroxide
B
The BEST fixative for blood smears is:
a) Bouin solution
b) Carnoy solution
c) B-5 solution
d) methanol
D
When used as a secondary fixative, osmium tetroxide should be:
a) used after lead citrate
b) heated prior to use
c) combined with alcohol
d) used under a chemical hood
D
Which of the following fixatives is recommended for use in lipid histochemistry?
a) Zenker solution
b) acetone
c) formalin-saline
d) calcium formalin
D
In the Cajal method for demonstrating astrocytes, sections of brain should be fixed in formalin that contains:
a) sodum acetate
b) ammonium bromide
c) mercuric chloride
d) calcium chloride
B
Hollande solution is a modification of which of the following fixatives?
a) Helly solution
b) Orth solution
c) Carnoy solution
d) Bouin slution
D
Tissue fixed in which of the following solutions must be posttreated for mercuric chloride pigment?
a) B-5
b) Zamboni
c) Carnoy
d) Orth
A
Acetone is recommended for the primary fixation of:
a) prostate tissue for IHC
b) kidney tissue for fluorescent Ab techniques
c) muscle tissue for enzyme histochemistry
d) brain tissue for the diagnosis of rabies
D
Fresh, unfixed tissue can be stored safely for a short time by:
a) keeping it in a freezer
b) wrapping it in saline-moistened gause and refrigerating it
c) placing it in physiologic saline at room temperature
d) leaving it in a dry specimen container on the counter with a note to the histologist
B
The nuclei in this image indicate: (pg 18)
a) excellent chromatin demonstration
b) the use of old hematoxylin
c) incomplete fixation
d) overhydration
C
One characteristic of Bouin solution is that it:
a) penetrates poorly
b) destroys delicate structures
c) mordants connective tissue stains
d) preserves erythrocytes
C
The fixation of tissue by physical methods can be accomplished by the use of:
a) microincineration
b) microwaves
c) frozen sections
d) alcohol
B
Fixation in Carnoy solution will result in:
a) swelling of the tissue
b) preservation of most cytoplasmic structures
c) superior staining of amyloid with Congo red
d) good preservation of red blood cells
C
The recommended fixative for tissue suspected of containing spirochetes is:
a) 10% neutral buffered formalin
b) Bouin solution
c) Zenker solution
d) Helly solution
A
Which of the following is frequently added to formalin solutions to help preserve immunoreactivity?
a) glycerin
b) sodium acetate
c) zinc salts
d) chromates
C
Which of the following fixatives is contraindicated when silver stains are to be done for Helicobacter pylori?
a) glutaraldehyde
b) neutral buffered formalin
c) glyoxal
d) Bouin solution
C
A fixative used for the preservation of some enzymes is:
a) Bouin solution
b) B-5 solution
c) acetone
d) isopropanol
C
A common reason for adding acetic acid to fixatives is to:
a)coagulate proteins
b) reduce shrinkage
c) preserve carbohydrates
d) change pH
B
Formaldehyde acts as a fixative by:
a) uncovering acid groups
b) coalgulating proteins
c) crosslinking proteins
d) rupturing peptide linkages
C
If it is known prior to fixation that a distinction must be made between collagen and muscle, the preferred fixative is:
a) neutral buffered formalin
b) Orth fluid
c) absolute alcohol
d) Bouin solution
D
Bouin fixation is contraindicated for Feulgen stains because during fixation:
a) nucleoproteins are precipitated
b) nuclei are excessively hydrolyzed
c) nucleoproteins are crosslinked
d) RNA is dissolved
B
When using Bouin fixative, the shrinking effect produced by one component is balanced by the swelling effect of:
a) formallin
b) acetic acid
c) osmium tetroxide
d) potassium dichromate
B
10% formalin is equivalent to what percent paraformaldehyde solution?
a) 40
b) 10
c) 4
d) 1
C
To obtain the same nuclear detail characteristic of cytologic preparations, which of the following percentage concentrations of isopropyl alcohol may be substituted for 95% ethyl alcohol?
a) 70
b) 80
c) 90
d)100
B
Which of the following renders fat insoluble for subsequent processing?
a) picric acid
b) osmium tetroxide
c) chloroform
d) formaldehyde
B
The presence of acetic acid in fixatives produces:
a) cell shrinkage
b) red cell destruction
c) protein coagulation
d) lipid preservation
B
Microscopic review of a formalin fixed tissue section demonstrates a fine, brown-black artifactual pigment. The artifact most likely could have been prevented by:
a) placing the tissue in formalin immediately after removal
b) preparing the solution just before use
c) washing the tissue after fixation
d) making the solution neutral
D
When osmium tetroxide is used as a fixative for paraffin embedding, it:
a) must be used after a primary fixative
b) should be used on very thin sections
c) should be made hypotonic to tissue
d) must be made basic
B
Which of the following fixative components produces shrinkage and penetrates poorly:
a) glacial acetic acid
b) mercuric chloride
c) potassium dichromate
d) ethanol
B
The component in Bouin solution that causes tissue shrinkage is:
a) picric acid
b) acetic acid
c) mercuric chloride
d) potassium dichromate
A
Which of the following fixatives should be selected when it is desirable to preserve erythrocytes in tissue?
a) Clark
b) Bouin
c) Carnoy
d) B-5
D
A tissue section reveals a dark brown microcrystalline pigment which is bireringent. To remove this pigment, the section should be treated with an alcoholic solution saturated with:
a) sodium thiosulfate
b) oxalic acid
c) picric acid
d) hydrochloric acid
C
Shrinkage and distortion of tissue is greatest following fixation in:
a) Bouin solution
b) Zamboni solution
c) absolute alcohol
d) 10% neutral buffered formalin
C
Tissue to be stained by the Warthin-Starry technique should be fixed in:
a) Zenker fluid
b) saturated mercuric chloride
c) formalin
d) osmium tetroxide
C
Because of the location of the pigment, it is most likely: (pg 22)
a) formalin
b) mercury
c) chromate
d) melanin
A
The tissue shown is: (pg 22)
a) esophagus
b) stomach
c) small intestin
d) colon
C
When fat needs to be preserved, the fixative of choice is:
a) formalin
b) Zenker solution
c) Carny solution
d) Bouin solution
A
Picric acid was used alone as a fixative for a section of liver. The tissue most likely will show:
a) extreme swelling
b) excessive hardening
c) decreased uptake of eosin
d) hydrolyzed nucleic acid
D
The primary purpose of fixation is the:
a) preservation of carbohydrates
b) coagulation of lipids
c) removal of tissue fluids
d) stabilization of proteins
D
Clark solution lyses erythrocytes because it contains:
a) formalin
b) picric acid
c) acetic acid
d) chloroform
C
The tissue component most affected by the problem shown in the image below is called the: (pg 23)
a) lamina propria
b) muscularis
c) epithelium
d) adventitia
C
Microscopic examination of a PAS stained section reveals marked nonspecific staining. This could be caused by fixation in:
a) Bouin solution
b) Gendre solution
c) 10% neutral buffered formalin
d) glutaraldehyde solution
D
The pathologist is sure that urate crystals were present in a tissue biopsy and should therefore be present in the H&E, but polarization of the tissue is negative. This could possibly result from the:
a) application of the wrong stain for demonstration
b) poor clearing & infiltration
c) fact that urate crystals are not birefringent
d) use of a water-based fixative
D
Formaldehyde reacts primarily with which of the following protein groups?
a) COOH
b) C=O
c) NH2
d) N=N
C
During embedding, white deposit is noted on several tissues. The tissues had been fixed in zinc-formalin, transferred to phosphate-buffered formalin, dehydrated with 65%, 95%, and absolute alcohols, and cleared with xylene. What might have caused the deposit?
a) left in zinc-formalin too long
b) fixed in incompatible fixatives
c) not washed properly
d) not dehydrated properly
C
Electron micrographs of a tissue section reveal electron-lucent membranes. This most likely indicates:
a) fixation was done in Bouin
b) fixative osmolality was wrong
c) specimen was not postfixed in osmium tetroxide
d) sections were stained with uranyl acetate
C
Michel medium is:
a) used for transporting unfixed tissue
b) indicated for long-term storage of fixed tissue
c) a fixative with limited use
d) an antigen retrieval solution
A
H&E evaluation from a surgically removed, formalin fixed sm bowel specimen shows an absence of much of the epithelium in otherwise normal tissue. Prevention?
a) opening the specimen and adding fixative upon receipt
b) avoid forcep use for dissection
c) fix specimens for 48hrs
d) increase formalin % in solution
A
A lab’s primary fixative has been changed from neutral buffered formalin to alcohol-based fixative promoted for antigen preservation. H&E show marked increase in eosin uptake. The new fixative is:
a) creating a different tissue isoelectric point
b) blocking protein precipitation
c) generating more neg charges
d) increasing corsslinking of the amino groups
A
H&E of small intestine in 10% NBF shows a complete absence of surface epithelium & poor cellular detail in the lamina propria. This is most likely the result of:
a) autolysis
b) mechanical trauma
c) antemortem changes
d) prolonged fixation
A
The problem seen on the section below is most frequently seen in which of the following specimens: (pg 25)
a) colonoscopy
b) post-mortem
c) surgical removal
d) edoscopic biopsy
B
When sucrose is used to treat tissue for enzyme histochemical studies:
a) cell membranes are mobilized
b) frozen sectioning is very difficult
c) the tissue must be removed after a brief period of treatment
d) the solutions should contain 30% sucrose and 1% gum acacia
D
Kidney tissue has been submitted in Michel medium for immunofluorescence studies. Before the HTL performs the required studies, the tissue should be:
a) washed in PBS containing 10% sucrose
b) frozen immediately upon receipt
c) placed in an aldehyde fixative
d) refrigerated
A
An isotonic solution for human tissue has an osmolality of approx:
a) 500 milliosmols
b) 340 milliosmols
c) 260 milliosmols
d) 150 milliosmols
B
Marked distortion of architecture and predominately pyknotic nuclei are noted on H&E of kidney. The tissue was fixed in the microwave in a saline solution. This problem could be prevented by:
a) changing the processing sched
b) using another fixative solution
c) carefully controling the temp
d) using longer fixation times
C
Bouin fixed tissue in paraffin blocks are taken from storage after several years. Newly cut H&E show no nuclear staining even though nuclei stained well prior to storage. To prevent this in the future, one must:
a) neutralize picric acid before processing
b) ensure Bouin solution is at neutrality
c) ensure formalin doesn’t contain formic acid
d) store blocks below 20C
A
H&E of formalin fixed specimen shows nuclear bubbling. To avoid in the future:
a) extend fixation time
b) fix at regrigerator temperatures
c) change the pH of the fixative
d) decrease osmolality of fixative
A
During electron microscopy print exam of normal kidney, disruption of the usual cellular morphology is noted. Possible cause is that the primary fixation occurred on osmium tetroxide for:
a) 1 minute
b) 30 minutes
c) 1 hour
d) 12 hours
D
Which of the following fixatives is stable at room temperature?
a) Karnovsky paraformaldehyde-glutaraldehyde
b) 2-3% glutaraldehyde
c) Zamboni PAF
d) 1% buffered osmium tetroxide
C
The paraformaldehyde in Zamboni PAF solution must be heated to 60C prior to adding 1M NaOH in order to:
a) dissociate paraformaldehyde
b) purify the glutaraldehyde
c) polymerize the gutaraldehyde
d) eliminate the gasses present
A
Prior to processing, tissue fixed in glutaraldehyde for electron microscopy should be:
a) postfixed in formaldehyde
b) rinsed w/ gum sucrose solution
c) postfixed in osmium tetroxide solution
d) rinsed with alcohol buffer solution
C
Poor fixation of electron microscopy specimens is often indicated by the appearance of:
a) evenly dispersed ground substance
b) stabilized cytoplasmic proteins
c) disrupted mitochondrial mumbranes
d) unaltered nuclear and cytoplasmic membranes
C
Formalin pigment may be removed from microscopic sections by treating them with:
a) Lugol iodine and sodium thiosulfate
b) potassium permanganate
c) dilute acetic acid
d) potassium hydroxide & 70% alcohol
D
Fixatives that tend to mask antigenic sites and hamper IHC localization of antigens contain:
a) mercury
b) phosphates
c) aldehydes
d) alcohol
C
Fallopian tube H&E reveals very muddy appearing nuclei with no visible chromatin pattern. This is most likely the result of fixation that was?
a) incomplete
b) delayed
c) prolonged
d) appropriate
A
In immuno-electron microscopy (IEM), a fixative other than osmium tetroxide is used because osmiu would:
a) blacken the tissue
b) potassium dichromate
c) destroy the antigenicity
d) decrease membrane preservation
C
The difference between Orth & Helly solutions is that ONLY Helly contains:
a) formaldehyde
b) potassium dichromate
c) mercuric chloride
d) acetic acid
C
A small biopsy is submitted with a request for “stat” acid-fast and fungus stains. The tissue should be fixed in:
a) Zenker solution
b) Carnoy fluid
c) B-5 fluid
d) buffered formalin
D
A microwave oven can be used for fixation because it:
a) causes crosslinking of proteins
b) induces physical fixation
c) increases tissue basophilia
d) inactivates enzymes with beta radiation
B
Unstained slides from B-5-fixed tissue should be treated with which of the following before staining?
a) lithium carbonate
b) potassium iodid
c) sodium bisulfite
d) iodine
D
Brown pigment is seen on top of the tissue. Adjecent sections are treated with: 1. iodine & sodium thiosulfate 2. potassium ferrocyanide & hydrochloric acid 3. saturated alcoholic solution or picric acid. All sections still show the brown pigment. this pigment could have resulted from improper washing following fixation in:
a) Zenker fluid
b) formalin
c) B-5 fluid
d) Bouin fluid
A
A possible rhabdomyosarcoma has been fixed in 10% NBF. For the nonIHC stains needed to confirm this tumor, microscopic sections should be:
a) prepared as frozen sections
b) treated with acetone before staining
c) mordanted in another fixative
d) deparaffinized with xylene-peanut oil mixture
C
Examination of the image below reveals a problem that most likely could have been prevented by:
a) washing with running water after fixation
b) neutralizing the tissue before processing
c) ensuring the use of buffered formalin
d) processed on a short cycle
C
When osmiom tetroxide is used on a kidney biopsy for ultrastructural studies, the time of fixation:
a) is not critical
b) should be 1hr or less
c) is prolonged b/c of tissue density
d) depends on the fixative concentration
B
Helly fixative prepared 3 weeks previously is now discoloed and turbid. The most likely reason for this is:
a) the presence of acetic acid in the fixative
b) improper solution buffering
c) the premature addition of formaldehyde
d) chemical oxidation
C
A section shows marked lysis of erythrocytes & the presence of brown crystalline pigment. The results indicate that the fixative was most likely:
a) glutaraldehyde
b) glyoxal
c) acetic acid-formalin
d) methacarn
C
A small amount of white precipitate is noticed in the bottom of the lab’s stock 37-40% formaldehyde container. the most appropriate action is to:
a) discard the solution
b) disregard the precipitate
c) acidify the solution slightly
d) add methanol to the solution
B
Acetone can be used to fix tissue for the demonstration of:
a) myelin sheaths
b) phospholipids
c) oxidoreductases
d) cell surface antigens
D
An unfixed hemorrhagic spleen tissue block is fixed for 24hrs in formalin that was prepared 6 months previously. Subsequent H&Es show a granular black pigment on the surface of the tissue. This problem can be prevented by:
a) wash tissue prior to fixation
b) ensure formalin is buffered
c) use iodine solution on processor
d) use only freshly made solutions
B
If tissue is fixed in formali, postfixation is critical for a nonIHC special stain used in the diagnosis of which of the following tumors:
a) liposarcoma
b) fibrosarcoma
c) neurosarcoma
d) rhabdomyosarcoma
D
The problem shown below most likely could have been prevented by: (pg 31)
a) more careful grossing technique
b) using different fixative solution
c) immediate contact with fixative
d) decreasing the flotation bath temp
C
Ago uty tophus is seen in a fresh tissue specimen. The ideal fixative for the crystals which may be occurring with this condition is:
a) buffered formalin
b) Bouin solution
c) absolute alcohol
d) Orth fluid
C
To ensure adequate fixation, a colon polyp measuring approx 2x2x3 cm should be placed in what volume of fixative, minimum?
a) 2.4mL
b) 24mL
c) 240mL
d) 2400mL
C
A characteristic of acetone fixation is that:
a) glycogen is removed from tissue
b) enzymes are removed from tissue
c) tissues are overhardened
d) antigen-antibody reactions are destroyed
C
A wet tissue sample has been requested for electron microscopy. All available tissue has been fixed in Bouin, washed, and stored in 70% ethanol. The tissue will show:
a) good fixation of nucleoprotiens
b) well preserved organelles
c) some shrinkage of nucleolus
d) inadequately preserved ultrastructures
D
A surgical specimen presumed to be metastatic malignant melanoma should be fixed in reagent compatible with the use of which of the following special stains or diagnostic procedures?
a) colloidal iron
b) IHC
c) immunofluorescence
d) digestion procedures
B
A section of tendon sheath has been fixed in 10% NBF for 24hrs, routinely processed, and embedded in paraffin. The demonstration of urate crystals:
a) should be performed after postfixation
b) will be best with fluorescence technique
c) must be performed on air-dried sections
d) will not be possible due to improper fixation
D
Clark solution differs from Carnoy solution in that only Carnoy contains:
a) absolute alcohol
b) acetic acid
c) chloroform
d) glyoxal
C
A section of healthy adrenal gland to be used as a control for chromaffin granules is fixed in osmium tetroxide for 8hrs. When examined, no granules are demonstrated. Resolve in the future by:
a) washing tissue before staining
b) select another tissue for control
c) use another fixative
d) increasing fixation time
C
An aqueous fixative MUST be chosen for sections that require:
a) the preservation of lipids
b) rapid sample fixation
c) maximum hardening
d) preservation of amyloid
A
for HER2 testing, it is recommended that fixation be no less than:
a) 2 hrs
b) 6 hrs
c) 12 hrs
d) 48 hrs
B
Tissues that undergo the most rapid autolysis are rich in:
a) connective tissue
b) enzymes
c) blood supply
d) bacteria
B
Pap smears that have been fixed with a spray fixative stained without another treatment. The most likely result will be:
a) excellent cytologic detail
b) the loss of cells from the slide
c) cells showing air-drying artifact
d) nuclei that appear foggy & lack detail
D
The problem shown most likely can be prevented by: (pg 33)
a) cutting the specimen in small slices and placing in fixative immediately
b) handling the sections more carefully during microtomy
c) decrease fixation time
A
All of the following are steps in the gluteraldehyde fixation procedure for electron microscopy specimens EXCEPT:
a) initial fixation in phosphate buffered glutaraldehyde
b) rinse with phosphate buffer following fixation
c) rinse with gum sucrose following the buffer wash
d) postfixation in osmium tetroxide
C
The BEST Ab for determining when tissue has been overfixed is:
a) S-100
b) CD20
c) vimentin
d) Cam 5.2
C
For HER2 Ab testing, it is recommended that the fixative be:
a) acetone
b) formalin
c) Bouin
d) none
B
Tissue antigens are irreversibly blocked by fixation in:
a) B-5
b) acetic zinc formalin
c) Bouin
d) glutaraldehyde
D
Smears stained by the Papanicolaou method show impaired nuclear staining. One possible explanation is that:
a) Gill II hematoxylin was used
b) smears were allowed to air dry
c) Bismarck brown was too conc.
d) sections were spray-fixed while wet
B
The nuclear problem seen could have been prevented by fixation that was:
a) decreased
b) prolonged
c) immediate
d) delayed
B
Which of the following fixatives is BEST when used at approx neutrality?
a) glyoxal
b) formalin
c) Bouin
d) Zenker
B
Relatively unnoted formalin pigment by light microscopy may be visualized if the slide is examined with which microscope?
a) trensmission election
b) dark field
c) polarizing
d) fluorescence
C
Pathologists are not happy with staining results after the fixative solution was changed. This is probably because the stain was not adjusted to account for the difference in:
a) penetration
b) cell shrinkage
c) fixative binding sites
d) enzyme preservation
C
Many aqueous fixatives penetrate poorly because of:
a) enzyme activity
b) fatty cell membranes
c) conformational changes
d) pH
B
Formalin pigment has been classified by some authors as:
a) endogenous lipidic
b) endogenous hematogenous
c) endogenous nonhematogenous
d) exogenous metallic
B
