1 : Fixation

An example of an additive fixative is one that contains:

a. picric acid

b. acetic acid

c. ethyl alcohol

d. acetone

a. picric acid

Although picric acid is considered an additive fixative, its reaction with proteins is not completely understood.

When compared with tissue fixed in formalin, tissue fixed in zinc-formalin will show:

a. better ultrastructural preservation

b. decreased immunize activity

c. increased enzyme activity

d. superior nuclear detail

d. superior nuclear detail

Zinc is less toxic than mercury. It has been substituted for mercury 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. microwaved fixation

d. frozen sectioning

a. incomplete fixation

If tissue is incompletely fixed before placing in the dehydrating solution, a nuclear bubbling artifact may result.

Microscopic examination 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. delayed fixation

When a 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. Michel solution

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. Hollande solution

The cupric acetate present in Hollande solution stabilized RBC membranes, so that the lysis that occurs in Bouin solution, and other acetic containing fixatives, is much less.

When the microwave oven is used for fixation, the most critical factor is:

a. preparation of the formalin solution

b. use of glass containers

c. control of the temperature

d. osmolality of the fixative solution

c. control of the temperature

Irreversible morphologic damage will result is the temperature is not carefully controlled.

To adequately remove the calcium from a specimen containing areas of microcalcification, the tissue should be fixed in:

a. Hollande solution

b. neutral buffered formalin

c. B-5 solution

d. Zamboni solution

a. Hollande solution

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

Hollande solution contains cupric acetate, which will stabilize RBC membranes and the granules of eosinophils and endocrine cells.

A specimen is submitted with the statement that it was fixed in formalin. Microscopic sections show marked lysis of erythrocytes. This indicates that the fixative was most likely:

a. prepared with too much formalin

b. buffered above neutrality

c. acidified with acetic acid

d. not formalin

c. acidified with acetic acid

Marked lysis of erythrocytes is a characteristic of fixatives containing acetic acid.

Fixatives are classified as additive because of the:

a. addition of several chemicals to the solution

b. additional, or binding of the fixative to the tissue proteins

c. additional reactions occurring with longer fixation

d. additional reactive tissue sites available for dye binding

b. addition, or binding of the fixative to tissue proteins

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. nonspecific PAS staining

Glutaraldehyde is a dialdehyde, and the extra aldehyde group is not involved in most crosslinking reactions; therefore, it is left free to react in any method using Schiff reactant, such as the acid-Schiff (PAS) stain. This leads to false positive results.

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. absolute alcohol

Uric acid crystals are water soluble, so they 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

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 are 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 metabolic processes

c. protect tissue against alteration during subsequent processing

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. retard the polymerization of formaldehyde

Commercial formalin contains about 10-14% methanol which is added to prevent polymerization of paraformaldehyde, a highly polymeric form of formaldehyde.

The problem shown in this image is the result of:

a. incomplete fixation

b. excessive dehydration

c. incomplete clearing

d. poor paraffin infiltration

a. incomplete fixation

The cracks in the tissue and the smudgy nuclei are due to incomplete fixation.

In electron microscopy, Zamboni fluid, glutaraldehyde, and osmium textroxide function as:

a. dehydrating elements

b. clearing agents

c. embedding media

d. fixative solutions

d. fixative solutions

These three solutions function as fixatives for specimens of electron microscopy. Osmium tetroxide may be used as either a primary of 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

Sodium borate is not a fixative.

Bouin solution is contraindicated for:

a. small tissue biopsies

b. tissue intended for subsequent trichrome stains

c. tissues to be stained by Fuelgan reaction

d. routine tissue sections

c. tissue to be stained by Fuelgan reaction

Picric acid is a sufficiently strong acid to hydrolyze nuclei, so if stains for DNA (Fuelgan) 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. ammonium hydroxide in 70% alcohol

100 mL of 70% alcohol containing 3 mL of ammonium hydroxide will remove formalin pigment when slides are placed in this solution for 30 minutes to 3 hours.

Stock neutralized formalin is prepared in the laboratory by storing the solution over a layer of calcium carbonate. The solution withdrawn from this stock solution will:

a. become acidic

b. become alkaline

c. remain neutral

d. exhibit metachromasia

a. become acidic

The solution 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

b. improper sectioning

c. poor staining

d. incorrect section placement

a. poor fixation

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 300 mL of 37% to 40% formaldehyde solution?

a. 1,800

b. 2,500

c. 2,700

d. 3,600

c. 2,700

300 mL of a 37-40% formaldehyde solution added to 2,700 mL of water will yield a total of 3,000 mL of a 10% formalin solution.

One action of acetic acid is to:

a. exert a shrinking effect on the tissue

b. render nucleoprotein acidophilic

c. form salt linkages between protein chains

d. coagulate nucleoproteins

d. coagulate nucleoproteins

The major use of acetic acid in fixatives is the precipitation, or coagulation, and preservation of nucleoproteins.

Aldehyde fixatives are used for electron microscopy preparations because they:

a. are readily available

b. visibly stained tissue

c. preserve cell infrastructure

d. coagulate tissue lipids

c. preserve cell infrastructure

Aldehyde fixatives are used for electron microscopy preparations because they preserve cell ultrastructure. They 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. will make tissue more receptive to eosin staining

Tissue that has been fixed in a solution containing potassium dichromate will be very receptive to eosin staining.

If mercuric chloride is used alone for fixation, it will:

a. leave tissue proteins in coagulated

b. produce a very acidic solution

c. penetrate poorly and cause excessive shrinkage

d. decrease tissue affinity for stains

c. penetrate poorly and cause excessive shrinkage

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 buffered 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 microscopic section with:

a. saturated alcoholic picric acid

b. alcoholic lithium chloride

c. iodine and sodium thiosulfate

d. potassium permanganate and oxalic acid

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 buffered formalin solutions. The pigment may be removed by treating in alcoholic picric acid.

For good fixation of tissue with osmium tetroxide for electron microscopy, it is recommended that the tissue segment be no longer than:

a. 1 mm³

b. 2 mm³

c. 1 cm³

d. 2 cm³

a. 1 mm³

Osmium tetroxide penetrates very poorly, and so specimens should be minced to approximately 1 mm cubes for electron microscopy and cut very thin for demonstrating fat in paraffin sections.

The tissue shown in this image is:

a. liver

b. kidney

c. spleen

d. lymph node

c. spleen

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. 50-70% alcohol

Traditionally, tissue fixed in Bouin solution is washed with 50-70% alcohol, or 70% alcohol saturated 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. formalin

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:

a. cell shrinkage

b. smudgy nuclei

c. pyknotic nuclei

d. nuclear bubbling

d. nuclear bubbling

Absolute ethanol is a poor choice for fixation of:

a. glycogen

b. pigments

c. lipids

d. blood smears

c. lipids

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

Glutaraldehyde is a dialdehyde. The extra aldehyde group does not form cross links with the tissue, leaving one aldehyde group free to react in techniques depending on the demonstration of aldehydes.

It is necessary to adjust the pH of most formalin solutions because if the presence of:

a. methanol

b. formic acid

c. paraformaldehyde

d. carbon dioxide

b. formic acid

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 approximately 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. temperature of the fixative solution

In general, an increase in temperature increases the rate of fixation but also increases the rate of autolysis and diffusion of cellular elements.

Carnoy solution is recommended for the preservation of:

a. acid-fast bacilli

b. nucleic acids

c. lipids

d. red blood cells

b. nucleic acids

Carnot solution exhibits good nuclear preservation, but lyses RBCs, 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. pH

Good preservation of tissue for light microscopy is least dependent on fixative pH, and many fixatives are quite acidic. Varying the pH from 4-9 apparently makes little difference in the fine structure produced by formalin fixation; however, a pigment is produced at the lower pH.

Formalin pigment is generally created in tissues fixed in formalin when the pH:

a. rises above 6

b. falls below 6

c. is buffered to neutrality

d. is 7.2

b. falls below 6

Formalin pigment is generally created in tissues fixed in formalin when the pH falls below 6. This pigment may be formed by any acidic fixative containing formaldehyde.

Very bloody cytology smears are often treated with:

a. 10% formalin

b. Hollande solution

c. Clark solution

d. acetone

c. Clark solution

Because blood sometimes obscures important cellular detail in very bloody cytology smears, Clark solution is used to lyse the red blood cells.

Glycol 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. rapidity of action

Glyoxol fixatives are extremely rapid in action, and surgical specimens are fixed after only 4-6 hours of exposure. Glyoxol forms cross links only under very specific conditions, lyses erythrocytes, and there may be a slight reduction in staining after long periods of storing.

If a tissue section 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 biosulfite

c. postfixed in the appropriate fixative prior to staining

d. treated with hydrogen peroxide

c. postfixed in the appropriate fixative prior to staining

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:

a. understained chromatin

b. cell shrinkage

c. smudgy nuclei

d. nuclear bubbling

c. smudgy nuclei

Fixation in Bouin solution is:

a. recommended for the Fuelgan reaction

b. excellent for ultrastructural preservation

c. the cause of considerable swelling of tissue

d. frequently used for endocrine tissues

d. frequently used for endocrine tissues

Bouin solution is an excellent fixative for biopsy specimens of the gastrointestinal tract 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 dichromate, and 37-40% formaldehyde

c. mercuric chloride, sodium acetate, and 37-40% formaldehyde

Pigment caused by mercury-containing fixatives can be removed from tissue by:

a. saturated alcoholic picric acid

b. iodine-sodium thiosulfate

c. washing in running water

d. potassium hydroxide in water

b. iodine-sodium thiosulfate

Iodine followed by sodium thiosulfate is used for the removal of the pigment caused by mercury containing fixatives.

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

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 enzymatic activity

b. permit the dehydration to function properly

c. inhibit crosslinking of tissue proteins

d. stabilize tissue carbohydrates

a. prevent decomposition due to enzymatic activity

Decomposition of tissue by enzymatic activity happens as soon as the blood supply is interrupted: therefore, the tissue should be placed in fixative solution immediately after removal.

Calcium-formalin fixative is recommended for the best preservation and subsequent demonstration of:

a. glycogen

b. phospholipids

c. amyloid

d. estrogen receptors

b. phospholipids

Calcium formalin is recommended especially for the fixation and preservation of phospholipids in tissues. Phospholipids tend to take up water and extend their surface by growing outward in wormlike myelin forms; calcium ions have a dramatic effect in preventing this.

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. 15-20 times

Ultrastructural preservation will be very poor following fixation in:

a. Zamboni PAF

b. 2% buffered glutaraldehyde

c. osmium tetroxide

d. 10% aqueous formalin

d. 10% aqueous formalin

Ultrastructural preservation will be poor following fixation in 10% aqueous formalin. Formalin solutions should be buffered to neutrality and the tonicity adjusted for use as a fixative for electron microscopy.

Glyoxol is a/an:

a. aliphatic hydrocarbon

b. aromatic hydrocarbon

c. dialdehyde

d. ketone

c. dialdehyde

Glyoxol is the smallest dialdehyde.

Zinc-formalin fixatives:

a. give poor infrastructural preservation

b. can be used to preserve enzymes

c. result in poor nuclear detail

d. will not coagulate tissue proteins

a. give poor infrastructural preservation

Zinc is a protein coagulant and the fixatives used for ultrastructural studies are noncoagulants.

Zamboni PAF refers to a fixative containing:

a. potassium dichromate, acetic acid, and formaldehyde

b. potassium aluminum sulfate and paraformaldehyde

c. buffered picric acid and formaldehyde

d. picric acid, acetic acid, and formaldehyde

c. buffered picric acid and formaldehyde

Zamboni refers to a buffered picric acid-formaldehyde solution that may be used as a general purpose fixative. It allows secondary fixation with osmium tetroxide and preserves the morphologic characteristics accurately; therefore, it is useful for both light and electron microscopy.

The problem seen in this image most likely could have been prevented by:

a. immediate and prolonged fixation

b. less processing time

c. increased paraffin infiltration

d. better microtomy technique

a. immediate and prolonged fixation

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. neutral buffered formalin

Stains for fat are done on frozen sections, and the preferred fixative is neutral buffered formalin.

Fixation of cytology smears should occur within:

a. 1-2 seconds

b. 10-15 seconds

c. 40-45 seconds

d. 1 minute

a. 1-2 seconds

Cytology smears should be fixed within 1-2 seconds, or an air drying artifact will most likely occur.

When osmium tetroxide is used as a fixative in histology, it:

a. destroys lipids

b. interferes with staining

c. leaves tissue very soft

d. distorts cell membranes

b. interferes with staining

After osmium tetroxide fixation, cell cytoplasm has little affinity for the anionic (acid, eg, eosin) dyes, but will readily accept cations (basic) dyes.

The breakdown of tissue due to enzymatic activity is called:

a. polymerization

b. putrefaction

c. autolysis

d. osmosis

c. autolysis

The breakdown of tissue due to enzyme activity is called autolysis. Putrefaction is caused by bacterial action.

The fixative of choice for the demonstration of gouty tophus is:

a. neutral buffered formalin

b. absolute alcohol

c. Bouin solution

d. Zenker solution

b. absolute alcohol

Urate crystals found in gouty tophi are water soluble; therefore, an aqueous based fixative cannot be used. Absolute alcohol is the fixative of choice.

A good fixative for routine use is one that:

a. makes tissue more permeable to fluids

b. is hypotonic to the tissue constituents

c. enhances putrefaction of tissue components

d. promotes tissue autolysis

a. makes tissue more permeable to fluids

A good fixative for routine use should make the tissue more permeable to fluids, so that all subsequent processes occur readily.

A pigment caused by chromate-containing fixatives can be prevented by treating the tissue prior to processing with:

a. running water

b. iodine

c. picric acid

d. potassium permanganate

a. running water

Chromate-containing fixatives should be washed with water before processing because the dehydrating alcohol can cause an insoluble pigment to form.

When fixing tissue for electron microscopy with formaldehyde or glutaraldehyd, the preservation of unrestricted depends on all of the following EXCEPT the:

a. pH

b. time and temperature

c. concentration and purity of the reagent

d. type of tissue

d. type of tissue

The pH, time and temperature, and the concentration and purity of the reagents all play an important role in the proper preservation of the ultrastructure. The type of tissue is not important.

Formic acid present in commercial formalin solutions may:

a. facilitate pigment formation

b. precipitate hemosideran

c. promote staining

d. cause tissue shrinkage

c. facilitate pigment formation

Formic acid in formalin leads to the formation of black acid hematin, or formalin pigment.

Carnoy solution is a combination of which of the following chemicals?

a. absolute alcohol, acetone, and glacial acetic acid

b. cedarwood oil, absolute alcohol, and glacial acetic acid

c. acetone, chloroform, and absolute alcohol

d. chloroform, glacial acetic acid, and absolute alcohol

d. chloroform, glacial acetic acid, and absolute alcohol

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. primary chromate fixative

A primary chromate fixative is necessary for the preservation of chromaffin granules. The demonstration of these granules are used for the diagnosis of pheochromocytoma.

When liver tissue is fixed with 2-3% glutaraldehyde:

a. glycogen is dissolved

b. the penetration rate is very rapid

c. a chemical reaction occurs with the lipids

d. the ultrastructure is preserved

d. the ultrastructure is preserved

2-3% glutaraldehyde is an excellent fixative for the preservation of tissue ultrastructure. Secondary fixation with osmium tetroxide is necessary to chemically react with a preserve the lipids before processing for EM.

A poor fixative is characterized by:

a. the absence of swelling or shrinking of tissue

b. inactivation of tissue enzymes

c. slow tissue penetration

d. the absence of distortion of dissolution

c. slow tissue penetration

The problem seen in this image possibly could have been prevented by:

a. buffering the formaldehyde solution

b. immediate contact with fixative solution

c. grossing the specimen more carefully

d. better processing protocol

b. immediate contact with fixative solution

Some of the epithelium is gone in this section, indicating that there was a slight delay in fixation.

Bouin solution contains all of the following except:

a. picric acid

b. absolute alcohol

c. 37-40% formaldehyde

d. glacial acetic acid

b. absolute alcohol

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 tissues

d. leave protein linkages unaffected

a. change the spongework of proteins into a mesh-like network

Coagulant fixatives establish a mesh-like network in tissue that allows solutions to readily penetrate or gain entry into the interior of the tissue.

The breakdown of tissue by bacterial action in called:

a. autolysis

b. putrefaction

c. denaturation

d. oxidation

b. putrefaction

When ultrastructure preservation is of the utmost importance, the fixative used should have a pH of:

a. 6.8-7.0

b. 7.2-7.4

c. 7.6-7.8

d. 8.0-8.2

b. 7.2-7.4

This is the physiologic pH, or approximately the pH of tissue fluid.

A fixative component that produces a brownish black pigment is:

a. picric acid

b. osmium tetroxide

c. mercuric chloride

d. acetic acid

c. mercuric chloride

Mercuric salts will produce a brownish-black pigment in tissues.

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. 70% ethanol

70% alcohol preserves both routine and immunohistochemical staining properties.

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. preserves glycogen very well

Ethyl alcohol is a precipitant fixative solution that is used primarily for the fixation of water soluble substances, such as glycogen and urate crystals. It is a nonadditive fixative that has no effect on tissue basophilia, and it shrinks and overhardens tissue.

The nuclear problem seen in this image is most often attributed to:

a. delay in fixation

b. incomplete fixation

c. overdehydration

d. poor paraffin infiltration

b. incomplete fixation

To prevent the formation of formalin pigment in tissues, formalin should be:

a. heated

b. cooled

c. buffered

d. acidified

c. buffered

Formalin solutions should be buffered to a pH of 6.8-7.4 in order to prevent the formation of formalin pigment; this pigment typically occurs if the pH drops below 6.0

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. 10% neutral buffered formalin

Specimens may remain in 10% neutral buffered formalin indefinitely

A biopsy that was placed in water by mistake is submitted to the laboratory. This mistake will most likely cause:

a. mushy sections

b. swollen and ruptured cells

c. hardening of the tissue

d. no appreciable changes

b. swollen and ruptured cells

Placing a specimen in water will cause the cells to take up water, swell, and rupture as a result. Placing tissue in any hypotonic solution can have this effect.

Sections of a breast carcinoma were fixed in a saline solution in the microwave oven. Microscopic examination of H&E stained sections show marked pyknotic, overstained nuclei. The staining results were most likely caused by:

a. use of saline for fixations

b. solution temperature exceeding 68*C

c. use of plastic containers in the microwave

d. presence of carcinoma in the breast tissue

b. solution temperature exceeding 68*C

If the temperature of the microwave oven is allowed to exceed 68*C, the tissue will show pyknotic, overstained nuclei.

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. hemosideran

d. mercury pigment

b. formalin pigment

An alcoholic solution saturated with picric acid is commonly used to remove formalin pigment; melanin pigment is usually bleached with an oxidizing agent: hemosideran, or iron pigment, is usually demonstrated with the Prussian blue reaction; and mercury pigment is removed by iodine.

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. causes reduction of some chemicals in the solution

Formaldehyde is a reducing agent, and will reduce some of the chemicals present in Helly solution. This results in the solution turning dark and turbid.

Formaldehyde solutions for routine use and most commonly buffered by:

a. monastic and dibasic phosphates

b. sodium acetate and acetic acid

c. s-colliding and hydrochloric acid

d. sodium barbitol and sodium hydroxide

a. monobasic and dibasic phosphates

Monobasic and dibasic sodium phosphates are most commonly used to buffer formaldehyde solutions.

The preferred fixative for the image seen below is:

a. 10% neutral buffered formalin

b. alcohol

c. Clark solution

d. Bouin solution

d. Bouin solution

The preferred fixative for the trichrome procedure shown in this image is Bouin solution.

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. may be used for electron microscopy

Zamboni fixative is a buffered formaldehyde-picric acid that is a good dual purpose fixative; that is, it is good for both light and electron microscopy.

The BEST fixative for blood smears is:

a. Bouin solution

b. Carnoy solution

c. B-5 solution

d. methanol

d. methanol

Methanol is the best fixative for blood smears.

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. used under a chemical hood

Osmium tetroxide vaporizes readily and the vapor itself readily fixed the nasal mucosa or the conjunctiva; contact with the vapor must be avoided.

Which of the following fixatives is recommended for use in lipid histochemistry?

a. Zenker solution

b. acetone

c. formalin-saline

d. calcium-formalin

d. calcium formalin

Calcium formalin is recommended for phospholipid preservation for any histochemical studies.

In the Cajal method for demonstrating astrocytes, sections of brain should be fixed in formalin that contains:

a. sodium acetate

b. ammonium bromide

c. mercuric chloride

d. calcium chloride

b. ammonium bromide

Formalin ammonium bromide is recommended for the fixation of tissue for staining with the Cajal method for astrocyte demonstration.

Hollande solution is a modification of which of the following fixatives?

a. Helly solution

b. Orth solution

c. Carnoy solution

d. Bouin solution

d. Bouin solution

Hollande solution is a medication of Bouin solution.

Tissue fixed in which of the following solutions must be post-treated for mercuric chloride pigment?

a. B-5

b. Zamboni

c. Carnoy

d. Orth

a. B-5

Tissue fixed in B-5 solution must be treated for the removal of the mercury pigment; B-5 solution contains mercuric chloride, sodium acetate, and formaldehyde.

Acetone is recommended for the primary fixation of:

a. prostate tissue for immunohistochemistry

b. kidney tissue for fluorescent antibody techniques

c. muscle tissue for enzyme histochemistry

d. brain tissue for the diagnosis of rabies

d. brain tissue for the diagnosis of rabies

acetone is recommended for frozen sections of brain tissue to be stained for diagnosis of rabies.

Fresh, unfixed tissue can be stored safely for a short time by:

a. keeping it in a freezer

b. wrapping it in a saline-moistened gauze and refrigerating it

c. placing it is a physiologic saline at room temperature

d. leaving it in a dry specimen container on the counter with a histologist

b. wrapping it in a saline-moistened gauze and refrigerating it

Fresh, unfixed tissue can be stored safely for a short time by wrapping it in saline-dampened gauze (excess saline squeezed out), and placing it on ice or in the refrigerator. It should never be placed in physiologic saline.

The nuclei in this image indicate:

a. excellent chromatin demonstration

b. the use of old hematoxylin

c. incomplete fixation

d. overhydration

c. incomplete fixation

The smudgy nuclei, or lack of chromatin definition, seen in this image is attributed to incomplete fixation of the tissue.

One characteristic of Bouin solution is that it:

a. penetrates poorly

b. destroys delicate structures

c. mordants connective tissue stains

d. preserves erythrocytes

c. mordants connective tissue stains

Bouin solution is the best fixative for connective tissue stains; if tissue is fixed in formalin, the sections should be mordanted in Bouin solution before staining with trichrome procedures. Bouin solution is excellent for preserving structures with soft and delicate textures, lyses red cells, and has good penetration.

The fixation of tissue by physical methods can be accomplished by the use of:

a. microincineration

b. microwaves

c. frozen sections

d. alcohol

b. microwaves

The heat generated by microwaves will physically fix tissue.

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. superior staining of amyloid with Congo red

Alcohol or Carnoy solution is preferred if Congo red staining for amyloid is to be done, although other fixatives may be used.