My Histo Tech Notes 1

Aug 2014

SECTION A: OBJECTIVE QUESTIONS (30 MARKS)

CHOOSE ONE BEST STATEMENT TO ANSWER THE QUESTION

1. What is “the science concerned with the microscopic structure of tissues and organs in relation to their function”?

(A) pathology

(B) histology*

(D) histological technique

2. What is “ to examine and dissect the surgical specimen”?

(A) fixation

(B) staining

(D) embedding

3. “To prevent tissue from spontaneous decay or prevent autolysis” is

(A) staining

(B) fixation*

(D) embedding

4. Dehydration is

(A) to introduce water to the tissue--hydration

(B) the last step in tissue processing--embedding

(D) applied in both tissue processing and staining procedure*

5. Decalcification process is to

(A) make the tissues looks better

(B) add calcium to the tissue for hardening

(D) increase the tissue chance to resist high temperature

6. Which of the following are criteria or Standard Operation Procedure for specimen receiving?

(A) unique identification number (MRN)*

(B) covered containers in buffered formalin

(D) All of the above

7. Below are the aims of fixation, EXCEPT

(A) retain shape or volume

(B) should not penetrate rapidly*

(D) to be as close as possible to their living state

prevent autolysis & putrefaction

8. What are the most used sections in laboratory classes, in order to protect the structures?

(A) cutting

(B) staining

(D) paraffin-embedded sections*

9. Which of the following fall under the category of simple fixatives?

(A) Formal saline*

(B) Carnoy’s fluid

(D) Potassium dichromate

10. What is the MOST COMMON staining method used?

(A) Wright’s

(B) Prussian blue

(D) Haematoxylin and eosin (H&E)*

11. What is the specimen needed for decalcification?

(A) skin

(B) bone*

(D) kidney

12. What is the fixative solution that can be used to preserve tissue specimen for long term storage?

(A) formal saline

(B) 10% formalin

(D) neutral buffered formalin*

13. What is the most commonly used dehydrating agent in tissue processing?

(A) petrol

(B) ethanol*

(D) methanol

14. Picking up section from floatation bath is usually called

(A) taking

(B) fishing*

(D) processing

15. Xylene is commonly used in which step of tissue processing?

(A) clearing*

(B) cleansing

(D) impregnation

16. What is the function of floating out ribbon in floatation bath?

(A) to produce tension in ribbon*

(B) to warm the tissue molecules

(D) to preserve protein in the cut tissue

17. What is the optimal thickness for tissue slices during sectioning?--2.5micron

(A) 0.8 mm

(B) 1-2 mm*

(D) 6-8 mm

18. Which of the following is NOT type of wax?

(A) ester

(B) resins

(D) capsicum*

19. Sudan dye stains for what component in the tissue?

(A) fat*--lipids

(B) sugar

(D) carbohydrate

20. What is the temperature inside the cryostat?

(A) 0°C

(B) 4°C

(D) -25°C*-- -15~-30

21. What is the aim of tissue processing?

(A) Kill any bacteria in the tissue prior to sectioning process

(B) Decrease the tissue size that will help for sectioning process

(D) To place tissue in a suitable support medium that will make easy for sectioning process*

22. Under which condition will a physician choose frozen section over paraffin section for a tissue specimen?

(A) Paraffin wax in the lab is out of stock.

(B) The histotechnologist lacks histopathology skill.

(D) The histotechnologist wants to shorten the processing time of the specimen.*

23. Xylene need to be applied again to the tissue slide at the end of the staining process because

(A) it will make the tissue slide free of dusts

(B) it kills the remaining bacteria in the tissue

(D) the slide needs to be cover-slipped using DPX which is permeable with xylene

24. Which of the following is NOT TRUE?

(A) Chemical should never be stored in alphabetical order.

(B) Storage outside the cabinet should be kept to a minimum.

(D) Glass containers should be stored on the bottom shelf of storage cabinet.

25. Statements below are the advantage of frozen sectioning, EXCEPT

(A) No staining required.*

(B) Productions are much more quickly than paraffin section.

(D) If more tissue is needed to make accurate diagnosis, surgeon is able to obtain additional sample and avoid a second operation.

26. All of the following statement describe the difference between cryostat and rotary microtome EXCEPT

(A) Cryostat has no knife while rotary microtome has it.*

(B) Cryostat is more expensive compared to rotary microtome.

(D) Cryostat require more skills and experiences to operate compared to rotary microtome that can be operated easily.

27. Histopathology laboratory may receive the specimens in the form of

I. biopsies

II. autopsies

III. whole organ

IV. body excretions

(A) I, IV

(B) III, IV

(D) I, III, IV

28. Which of the following equipments are used in sectioning process?

I. hot plate

II. microwave

III. floatation bath

IV. rotary microtome

(A) II, IV

(B) III, IV

(D) II, III, IV

29. What are the reagents contain in Harris’s Haematoxylin stain?--absolute alcohol, K alum, distilled water, glacial acetic acid

I. Vanette alkyl

II. Haematoxylin

III. Mercuric oxide

IV. Schiff’s reagent

(A) II, IV*

(B) II, III

(D) II, III, IV

30. Which of these following special stains are CORRECTLY matched?

STAIN: TARGET COMPONENTS

(A) Masson trichome iron--collagen, muscle, RBC

(B) Sudan black collagen--lipids, lipofuscins

(D) Verhoeff’s van gieson reticulum--muscle, collagen

SECTION B: SHORT ANSWER QUESTIONS (30 MARKS)

ANSWER ALL QUESTIONS

1. a. With examples each, list FIVE (5) types of specimen categories. (10 Marks)

2. a. State FOUR (4) methods of fixation. (4 Marks)

b. State SIX (6) factors involved with fixation. (6 Marks)

3. a. Give ONE (1) example of an acidic dye used in staining for histological slides? (1 Mark)

b. Give ONE (1) example of a basic dye used in staining for histological slides? (1 Mark)

c. Give THREE (3) examples of special stains and name the specific tissue/tissue components they stain? (3 Marks)

d. Give FIVE (5) factors affecting staining reaction? (5 Marks)

SECTION C: ESSAY QUESTIONS (40 MARKS)

ANSWER TWO (2) QUESTIONS ONLY

Question 1

Create a histopathology process flow. Your answers should include a brief discussion on each step of the process flow. (20 Marks)

Question 2

Discuss technical problems and troubleshooting histotechnologist encounters during fixation, processing, embedding and staining. (20 Marks)

Question 3

Compare rotary microtome and cryostat. Your answers must include the advantages and disadvantages. (20 Marks)

Aug 2013

SECTION A : OBJECTIVE QUESTIONS (30 MARKS)

Choose ONE best statement to answer the questions.

1. Histology laboratory may receive the specimens in the form of

I. smears

II. biopsies

III. autopsies

IV. whole organs

(A) I and II

(B) II and III

(D) II, III and IV*

2. Decalcification is a procedure to

(A) add calcium to the tissue

(B) make the tissue morphology clearer

(D) increase the chance of tissue to withstand the fixative solution

3. What are the purposes of tissue fixation?

I. to retain shape and volume of tissue

II. to preserve tissue substances and proteins

III. to allow autolysis and bacterial attack to the tissues--prevent

IV. to harden and protect the tissue from subsequent treatments

(A) I and II

(B) II and III

(D) II, III and IV

4. What is the MOST COMMON type of fixative used for paraffin section?

(A) Bouin’s fluid

(B) formaldehyde

(D) glutaraldehyde

5. What is the fixative solution that can be used for rapid fixation for urgent biopsy material and is not used for routine?

(A) formol saline

(B) 10% formalin

(D) Carnoy’s fixative

6. Factors involoved in tissue fixation are

I. pH

II. osmolarity

III. penetration rate

IV. time interval from removal to fixation

(A) I, II and III

(B) I, II and IV

(D) All of the above

7. The aim of tissue processing is to

(A) kill any microorganisms in the tissue

(B) remove all proteins in the tissue to aid in staining

(D) place tissue in a suitable support medium that will make the sectioning process easier*

8. Which of the below is in CORRECT order for tissue processing?

(A) dehydration, clearing, deparaffinization and bluing

(B) hydration, cleansing, deparaffinization and embedding

(D) dehydration, clearing, infiltration and impregnation, and embedding*

9. Dehydration

(A) is the last step in tissue processing

(B) is done by using descending grades of alcohol

(D) is applied both in tissue processing and staining procedures*

10. The MOST COMMON dehydrating agent used is

(A) petrol

(B) ethanol*

(D) methanol

11. The MOST COMMONLY used agent for removing alcohol in tissue processing is

(A) xylene*

(B) ethanol

(D) H&E stains

12. Paraffin wax as the embedding media is usually chosen because

(A) can be recycled

(B) white and looks nice

(D) cheap and large number of tissue blocks can be processed in a short time*

13. How long does the routine overnight automated tissue processing method take?--20hrs

(A) 8 hours

(B) 12 hours

(D) 25 hours

14. Which factor DOES NOT affect the rate of impregnation during tissue processing?

(A) heat

(B) noises*

(D) agitation

15. Rotary microtome, floatation bath and hot plate are used during

(A) fixation

(B) staining

(D) processing

16. The microtome blade should be

(A) heated up before use

(B) chilled before sectioning

(D) left exposed even when sectioning has finished

17. What is the function of floating out the ribbon in the floatation bath?

(A) to test if the ribbon can float or not.

(B) to warm the tissue section before staining

(D) to produce tension and spread out the ribbon*

18. The ideal temperature for the water in the water bath is 45

(A) the same as the melting point of paraffin wax--60

(B) 5ºC lower than the melting point of paraffin wax

(D) 10ºC higher than the melting point of paraffin wax

19. What is the advantage of having the water bath base black in color?

(A) cheaper

(B) able to absorb more heat

(D) temperature can be adjusted more quicker

20. Prolonged floating of ribbon in the water bath

(A) may expand the tissue and become distorted*

(B) allows oxidation of the molecules in the tissue

(D) can cause wrinkles and folds to develop in the tissue

21. Picking up the tissue section from the floatation bath is called

(A) taking

(B) mining

(D) pooling

22. Which is TRUE regarding trimming and sectioning the paraffin block?

(A) Thickness for trimming is 5 µm, whereas sectioning is 10 µm.

(B) Trimming needs to be carried out each time before sectioning is done.

(D) Paraffin block needs to be cooled before trimming but not necessary before sectioning.

23. Which of the following are FALSE of staining?

(A) Makes optical differentiation harder.*

(B) Accomplished by the use of dyes and stains.

(D) Sensitivity is the quality of successful staining which describes its ability to color one component and leaving others unstained.

24. Hematoxylin stains the

(A) nucleus*

(B) cytoplasm--eosin

(D) collagen fibers

25. Iron hematoxylin’s mordant is

(A) iron salt*

(B) aluminium

(D) phosphotungstic acid

26. Decolorization is associated with

(A) direct staining

(B) indirect staining

(D) progressive staining

27. The commonly used kind of Eosin is

(A) eosin Y*

(B) eosin B

(D) ethyl eosin

28. Which is TRUE regarding deparaffinization?

(A) uses 95% ethanol--xylene

(B) removal of wax is not necessary--removal with solvent is necessary

(D) makes stain permeate tissue section with highly organic solvents*

29. The removal of xylene during tissue staining is done by

(A) 10 dips in alcohol

(B) 1-2 changes of absolute alcohol*--1 min

(D) gradual run through decreasing concentrations of alcohol

30. This process facilitates easy handling and storage.

(A) clearing

(B) mounting*

(D) deparaffinization

SECTION B : SHORT ANSWER QUESTIONS (30 MARKS)

Answer ALL questions.

1. a) Classify the FIVE (5) types of chemical fixatives and give an example of each. (5 Marks)

b) Compare and contrast the mode of action of any TWO (2) types of chemical fixatives. (4 Marks)

c) Propose the type of fixation that can be used for substances which are soluble where chemical fixation is not possible. (1 Mark)

2. a) Give THREE (3) situations that require manual tissue processing. (3 Marks)

b) Briefly describe the steps for the embedding procedure using the moulds.(4 Marks)

c) Name THREE (3) types of embedding media. (3 Marks)

3. Provide the causes for the following problems in paraffin section.

a) Ribbon/consecutive sections curved. (2 Marks)

b) Splitting of sections. (3 Marks)

c) Sections roll into a tight coil. (3 Marks)

d) Sections expand & disintegrate on water surface (2 Marks)

PART C : ESSAY QUESTIONS (40 MARKS)

Answer TWO (2) questions only.

Question 1

Explain the SEVEN (7) main steps of tissue processing. (20 Marks)

Question 2

Write all steps in standard Haematoxylin and Eosin (H&E) staining procedure. (20 Marks)

Question 3

Explain in detail the factors involved in fixative preparation. (20 Marks)

Jan 2015

SECTION A: OBJECTIVE QUESTIONS (30 MARKS)

CHOOSE ONE BEST STATEMENT TO ANSWER THE QUESTIONS

1. Which of the following specimens is NOT to be processed in histologpathology laboratory?

(A) left ovary

(B) liver biopsy

(D) bloody sputum*

2. The MOST commonly used cross-linking fixative is

(A) ethanol

(B) picric acid

(D) Bouin’s solution

3. Which of the following refers to neutral buffered formalin (NBF)?

(A) It removes water from tissue.

(B) It coagulates several basic groups of proteins.

(D) It is highly miscible with clearing agent like xylene.

4. The addition of a few drops of 1% eosin to the specimen container prior to tissue processing procedure may

(A) improve the preservation of tissues

(B) kill any infectious agent present in the tissue

(D) increase the rate of tissue infiltration and impregnation with paraffin wax

5. The duration of bone decalcification using strong acids should be within

(A) 4 to 6 days

(B) 1 to 5 hours

(D) 24 to 48 hours*--if more than 24~48hrs, deterioration occurs

6. Which of the following is a dehydrating agent in tissue processing?

(A) toluene

(B) paraffin

(D) chloroform

7. The main purpose of tissue processing is to

(A) embed the tissue in a solid medium*

(B) introduce water into the tissue and cells

(D) preserve the tissue substances and proteins

8. What is the disadvantage when using xylene as a clearing agent?

(A) It is non-recyclable.

(B) It is less miscible混溶 with paraffin.

(D) It may cause over hardening of tissue if prolonged.*

9. The melting point of paraffin wax ranges from

(A) 20 to 40°C

(B) 40 to 70°C*

(D) 90 to 110°C

10. Choose the CORRECT steps of tissue processing:

(A) deparaffinization  hydration  clearing  embedding

(B) dehydration  clearing  deparaffinization  mounting

(D) dehydration  clearing  infiltration and impregnation  embedding*

11. What is the most suitable thickness for tissue block trimming?

(A) 1 cm

(B) 3 µm

(D) 200 mm

12. Which of the following processes involves the removal of xylene from tissues and replaced by paraffin wax?

(A) clearing

(B) infiltration*

(D) dehydration

13. The wax must be rapidly cooled on ice after embedding to

(A) prevent formation of bubbles

(B) minimize damage to the tissues

(D) preserve tissue proteins and lipids

Figure 1

To expose suitable area of tissue for sectioning

14. Which of the following processes is defined in Figure 1?

(A) fishing

(B) clearing

(D) embedding

15. Prolonged time on water during floating out sections will cause the tissue to

(A) dry

(B) shrink

(D) compress

16. Which of the following is useful to overcome hard tissue problem during sectioning procedure?

(A) re-orientate the tissue block

(B) re-embed the tissue in epoxy resin*

(D) expose the block surface to running tap water

17. What is the advantage of using an oven for drying of sections?

(A) Dust is less likely to settle on sections.*

(B) The sections do not need to be stained.

(D) Any microorganism in the sections will be killed.

18. The most suitable temperature for cutting non-fatty, unfixed tissues in frozen section is between

(A) -5 to -10°C

(B) -10 to -15°C

(D) -20 to -25°C*

19. Which of the following is NOT a form of dye-tissue interaction?

(A) hydrogen bonding

(B) colorimetric effect*

(D) Van der Waals’s forces

covalent--dye-tissue

hydrophobic effect--solvent-solvent

20. Harris’s haematoxylin is categorized as

(A) iron haematoxylin--Weigert's

(B) alum haematoxylin*

(D) haematoxylin without mordant

21. Deparaffinization step in haematoxylin and eosin staining procedure is carried out using

(A) xylene*

(B) DPX resin

(D) 0.5% acid alcohol

22. Which of the following combination is TRUE regarding tissue components and haematoxylin and eosin staining results?

(A) nuclei – grey--blue, black

(B) cytoplasm – pink*

(D) red blood cells – light blue--orange/red

collagen--pale pinky red

23. What is the medium commonly used in slide mounting / coverslipping?

(A) xylene

(B) DPX resin*

(D) immersion oil

24. Which of the following staining methods used to stain calcium?

(A) Oil Red O--lipids

(B) Von Kossa*

(D) Masson-Fontana--CT, melanin

25. The Congo Red staining technique is primarily useful in the diagnosis of

(A) amyloidosis*

(B) tuberculosis

(D) Alzheimer disease

26. Which of the following special stains is useful in the diagnosis of mesotheliomas?

(A) Oil Red O

(B) Von Kossa

(D) Masson-Fontana

27. Melanin in tissues can be stained using which staining method?

(A) Von Kossa

(B) Masson-Fontana*

(D) Gordon and Sweet’s--reticular fiber

28. The most widely used enzyme labels in immunohistochemistry (IHC) procedure are

I. RNA polymerase

II. alkaline phosphatase

III. horseradish peroxidase

IV. bacterial-derived β-D-galactosidase

(A) I and III

(B) I and IV

(D) III and IV

29. Heat induced epitope retrieval (HIER) step in immunohistochemistry may be carried out using

I. Incubator

II. Water bath

III. Microwave

IV. Pressure cooker

(A) I and II

(B) I and III

(D) III and IV

30. The aims of using vacuum in automated tissue processor is to

I. remove clearing agents

II. remove air bubbles in the tissue

III. increase the strength of tissue fibres

IV. remove abnormal proteins in the tissue

(A) I and II*

(B) I and III

(D) I, III and IV

SECTION B: SHORT ANSWER QUESTIONS (30 MARKS)

ANSWER ALL QUESTIONS

1. a. Define fixative. (2 Marks)

b. Provide the aims of fixation. (3 Marks)

c. Name TWO (2) categories of chemical fixatives and give an example for each. (2 Marks)

d. List SIX (6) factors that may affect the quality of fixation. (3 Marks)

2. a. Define tissue processing. (2 Marks)

b. Provide the aims of tissue processing. (3 Marks)

c. State and explain briefly FIVE (5) factors that affect the rate of tissue processing. (5 Marks)

3. a. Name the equipment used in production of section. (1 Mark)

b. Provide the causes and solutions for the following problems in production of sections.

i. Splitting of sections. (3 Marks)

ii. Sections do not join to form a ribbon. (3 Marks)

iii. Sections roll into a coil. (3 Marks)

SECTION C: ESSAY QUESTIONS (20 MARKS)

ANSWER TWO (2) QUESTIONS ONLY

Question 1

Describe the protocol of haematoxylin and eosin staining for paraffin sections. (10 Marks)

Question 2

Discuss briefly the Masson’s Trichrome staining in terms of uses, reagents and expected results. (10 Marks)

Question 3

Explain the Labeled StreptAvidin-Biotin (LSAB) method in immunohistochemistry. (10 Marks)

Jan 2014

SECTION A : OBJECTIVE QUESTIONS (30 MARKS)

Choose ONE best statement to answer the questions.

1. Which of the following is NOT a goal of tissue fixation?

(A) kill microorganisms

(B) freeze cell components*

(D) prevent enzymatic self-digestion

2. Which of the following fixation agents works well for electron microscopy and preserves lipoprotein membrane better?

(A) formaldehyde

(B) permanganate

(D) osmium tetroxide*

3. The combination of embedding and tissue sectioning technique which is best for making a 4 micron section would be

(A) ice; cryostat

(B) plastic; microtome

(D) paraffin; microtome*

4. Which imaging technique has the highest magnification?

(A) light microscope--2000X

(B) electron microscope*--1~5 MillionX

(D) fluorescence microscope--100X

5. Choose the INCORRECT statement about hematoxylin dyes.

(A) dyes DNA/RNA

(B) dyes mitochondria*

(D) usually associated with a mordant

6. Choose the INCORRECT statement about eosin dyes.

(A) dyes lysosome*

(B) dyes molecules pink

(D) produces good cytoplasmic staining

7. Which of the following is TRUE regarding iron hematoxylin?

(A) The mordant is aluminium.--iron salts

(B) Iron salts are ferric chloride.*

(D) Iron mordant stains the cytoplasm black.--violet black nuclei

8. The mordant in Harris’s hematoxylin is

(A) lead salts

(B) aluminium*--alum H

(D) phosphotungstic acid

9. Which of the following is MOST COMMONLY used to counterstain alum hematoxylin?

(A) eosin*

(B) safranin--gram staining

(D) Persian blue--iron--bright blue--anemia

10. The MOST COMMONLY used agent for removal of alcohol, making tissue transparent is

(A) xylene*--clearing

(B) ethanol

(D) paraffin

11. Clearing is done by

(A) 10 dips in alcohol

(B) 2-3 changes of xylene*

(D) gradual run through decreasing concentration of alcohol

12. The commonly used kind of eosin is

(A) eosin B

(B) eosin Y*

(D) Weigert’s eosin

13. Which of the following is TRUE regarding the mounting medium?

(A) water soluble

(B) liquid paraffin

(D) used for easy handling and storage of tissue sections on slides*

14. Choose the TRUE statement about deparaffinization.

(A) uses 95% ethanol--xylene

(B) 1-2 changes of alcohol for 1 minute

(D) makes stain permeate tissue section with water-based dyes*

15. Which type of fixative belongs to the picrate group?

(A) formalin

(B) dichromate

(D) osmium tetroxide

16. Which of the equipments below are NOT used during sectioning?

(A) coverslip

(B) hot plate

(D) floatation bath

17. The temperature for the floatation bath should be set

(A) 2°C below melting point of paraffin wax

(B) 2°C above melting point of paraffin wax

(D) 10°C below melting point of paraffin wax*

18. The ideal temperature for the water in the water bath is

(A) the same as the melting point of paraffin wax

(B) 5ºC lower than the melting point of paraffin wax

(D) 10ºC higher than the melting point of paraffin wax

19. Which of the following is the purpose of using a brush or forceps during sectioning?

(A) to curl the paraffin ribbon

(B) to remove folds from the ribbon*

(D) to make holes in between two adjacent tissue sections

20. Which of the following is NOT used as a section adhesive?

(A) raisin*--resin

(B) starch

(D) albumin

21. What are the requirements for a good paraffin ribbon?

(A) blunt blade

(B) microtome which is cooled

(D) tissue block which is fixed and embedded well*

22. Which is TRUE regarding trimming the paraffin block?

I. removal of extra wax

II. expose suitable area for sectioning

III. necessary to cool the blocks before trimming

IV. straight away produce the paraffin ribbon for fishing

(A) I and II*

(B) I and III

(D) III and IV

23. Which of the following is TRUE of tissue section thickness?

(A) during trimming, thickness is set at 10 µ

(B) to trim faster, thickness can be set at 20 µ*

(D) during sectioning of routine tissue blocks, 1.5 µ

24. What happens during the infiltration or the impregnation process of tissue processing?

(A) water flows out and alcohol flows in

(B) alcohol flows out and xylene flows in

(D) xylene flows out and paraffin wax flows in*

25. Which of the below should NOT be carried out to large surgical specimens that arrive in unfixed plastic bags while waiting for the pathologist to examine?

(A) put in fixative

(B) kept in refrigerator*

(D) wrapped with saline moistened towels

26. The appropriate volume of fixative recommended is

(A) 1-2 times the volume of tissue block

(B) 5-10 times the volume of tissue block

(D) 20-30 times the volume of tissue block

27. This is a fixative that can be used in vapour form.

(A) formaldehyde*

(B) Helly’s fixative

(D) osmium tetroxide

28. Which is the MOST COMMON agent used for dehydration?

(A) ethanol*

(B) formalin

(D) methanol

29. Which clearing agent is considered best for microscopy but cannot be used for routine work due to its high cost?

(A) toluene

(B) benzene

(D) cedar wood oil*

30. These factors increase the rate of impregnation EXCEPT

(A) decreasing the temperature*

(B) vacuum applied in wax jar

(D) smaller molecules reduce the viscosity

SECTION B : SHORT ANSWER QUESTIONS (30 MARKS)

Answer ALL questions.

1. a. Define autolysis and putrefaction. (2 Marks)

b. Briefly explain how autolysis and putrefaction is linked with fixation. (2 Marks)

c. Discuss THREE (3) physical factors that influence fixation. (6 Marks)

2. There are FIVE (5) groups of chemical fixatives.

a. TWO (2) fixatives are defined as crosslinking and precipitating. Discuss the difference in mechanism and give examples of both. (4 Marks)

b. Name the other THREE (3) groups of chemical fixatives and provide an example each. (6 Marks)

3. a. Which component of the cell stains blue with H&E stain? (1 Mark)

b. When viewing under the microscope, what are stained with eosin? (3 Marks)

c. Differentiate progressive staining and regressive staining technique. (2 Marks)

d. Compare direct staining with indirect staining. (2 Marks)

e. Dyes or reagents can enter tissues passively or actively. Explain. (2 Marks)

PART C : ESSAY QUESTIONS (40 MARKS)

Answer TWO (2) questions only.

Question 1

Explain the SEVEN (7) main steps to produce a tissue section. (20 Marks)

Question 2

Prepare an outline of the procedure for staining paraffin sections. (20 Marks)

Question 3

List the various faults in paraffin wax sectioning. Explain what are the causes and remedies for each fault. (20 Marks)

Jan 2013

SECTION A: OBJECTIVE QUESTIONS (30 MARKS)

Choose ONE best statement to answer the questions.

1. Which of the following specimens is NOT to be processed in histologpathology laboratory?

(A) left kidney

(B) lung biopsy

(D) bloody sputum*

2. The MOST commonly used cross-linkage fixative is

(A) alcohol

(B) picric acid

(D) osmium tetroxide

3. Which of the following is the advantage of neutral buffered formalin (NBF) fixative?

(A) It removes free water from tissue.

(B) It coagulates several basic groups of proteins.

(D) It is highly miscible with clearing agent like xylene.

4. The addition of a few drops of 1% eosin to the specimen container prior to tissue processing step can

(A) improve the preservation of tissues

(B) kill any infectious agent present in the tissue

(D) increase the rate of tissue impregnation with paraffin wax during tissue processing

5. The duration of bone decalcification using strong acids should be within

(A) 4 to 6 days

(B) 1 to 5 hours

(D) 24 to 48 hours*

6. Which of the following chemicals can be used as a dehydrating agent in tissue processing?

(A) paraffin

(B) toluene

(D) chloroform

7. The primary aim of tissue processing procedure is to

(A) embed the tissue in a solid medium*

(B) introduce water into the tissue and cells

(D) soften the tissue prior to the sectioning procedure

8. What is the disadvantage when using xylene as a clearing agent in tissue processing?

(A) It is non-recyclable.

(B) It is less miscible with paraffin.

(D) Over-exposure will cause over hardening.*

9. The melting points of paraffin wax ranges from

(A) 20 to 40°C.

(B) 40 to 70°C.*

(D) 90 to 110°C.

10. In sectioning procedure, tubular structures like arteries, fallopian tubes and vas deferens need to be cut in

(A) multiple orientations--multiple pieces with epithelial surface

(B) cross-section of the lumen*

(D) both transverse and longitudinal planes--muscle

skin biopsies, shave punch/excisions, cross section of epidermis, dermis, subcutaneous layer

11. Which of the following stage involves the removal of xylene from tissues and replaced by paraffin wax?

(A) Clearing

(B) Infiltration* & impregnation

(D) Dehydration

12. Which of the following tissues require specialized processing due to the delicate nature of the structures?

(A) skin

(B) eyes

(D) muscles

13. The wax must be rapidly cooled on ice after embedding to

(A) minimize damage to the tissues

(B) prevent the formation of bubbles--vacuum

(D) preserve the tissue proteins and lipids

Figure 1

To expose suitable area of tissue for sectioning

14. Which of the following processes is best described by Figure 1?

(A) fishing

(B) clearing

(D) embedding

15. Prolonged time on water during floating out sections will cause the tissue to

(A) dry

(B) shrink

(D) compress

16. Which of the following is useful to overcome hard tissues problem in sectioning procedure?

(A) Re-orientate the tissue block.

(B) Re-embed the tissue in epoxy resin.*

(D) Expose the block surface to running tap water.

17. What is the advantage of using an oven for drying of sections?

(A) Dust is less likely to settle on sections.*

(B) The sections do not need to be stained.

(D) Any microorganism in the sections will be killed.

18. The most suitable temperature for cutting non-fatty, unfixed tissues, in frozen section is between

(A) -5 to -10°C

(B) -10 to -15°C

(D) -20 to -25°C*

19. Which of the following is NOT a form of dye-tissue interaction?

(A) Hydrogen bonding

(B) Colorimetric effect*

(D) Van der Waals’s forces

20. Harris’s haematoxylin is categorized as

(A) iron haematoxylin

(B) alum haematoxylin*

(D) haematoxylin without mordant

21. What is the MOST commonly used chemical for the deparaffinization step in staining procedure?

(A) Xylene*

(B) DPX resin

(D) 0.5% acid alcohol

22. Which of the following combination is TRUE regarding tissue components and the Haematoxylin and Eosin staining results?

(A) Red blood cells – light blue--red/oren

(B) Muscle fibers – purple--pinky red

(D) Cytoplasm – pink*

23. Iron haematoxylins are better than alum haematoxylins because they

(A) have strong oxidizing ability

(B) use easy-to-prepare solutions*

(D) stain wider range of tissue structures

24. Which of the following staining method can be used to stain lipid?

(A) Oil Red O*

(B) Von Kossa

(D) Masson-Fontana

25. The Congo Red staining technique is primarily useful in the diagnosis of

(A) amyloidosis*

(B) tuberculosis

(D) Alzheimer disease

26. Which of the following special stains is useful in the diagnosis of mesotheliomas?

(A) Oil Red O

(B) Von Kossa

(D) Masson-Fontana

27. Melanin and argentaffin molecules in tissues can be stained using which staining method?

(A) Von Kossa

(B) Masson-Fontana*

(D) Gordon and Sweet’s

28. The most widely used enzyme labels in immunohistochemistry procedure is

(A) RNA polymerase

(B) alkaline phosphatase**

(D) bacterial-derived β-D-galactosidase

29. Positive expression of CD45 antigen in immunohistochemistry procedure may be associated with

(A) vascular tumours

(B) B-cell lymphoma*

(D) monocytic leukaemia

30. The aims of using vacuum in an automated tissue processor is to

I. remove clearing agents

II. remove air bubbles in the tissue

III. increase the strength of tissue fibres

IV. remove abnormal proteins in the tissue

(A) I and II*

(B) I and III

(D) I, III and IV

SECTION B: SHORT ANSWER QUESTIONS (30 MARKS)

Answer ALL questions.

1. a. Define fixative. (2 Marks)

b. Provide the aims of fixation. (3 Marks)

c. Name TWO (2) categories of chemical fixatives and give an example for each. (2 Marks)

d. List SIX (6) factors that may affect the quality of fixation. (3 Marks)

2. a. Define tissue processing. (2 Marks)

b. Provide the aims of tissue processing. (3 Marks)

c. State and explain briefly FIVE (5) factors that affect the rate of tissue processing. (5 Marks)

3. Provide the causes and solutions for the following problems in production of sections.

b. Ribbon or consecutive sections are curved. (4 Marks)

c. Splitting of sections. (3 Marks)

d. Sections roll into a tight coil. (3 Marks)

SECTION C: ESSAY QUESTIONS (40 MARKS)

Answer TWO (2) questions only.

Question 1

Explain the procedural outlines and protocol of Haematoxylin and Eosin (H&E) staining for paraffin sections. (20 Marks)

Question 2

Explain the procedure of Gomori’s silver impregnation method for reticular fibers. (20 Marks)

Question 3

Discuss the Labeled StreptAvidin-Biotin (LSAB) method in immunohistochemistry with example. (20 Marks)

Jan 2010

PART A – OBJECTIVE QUESTIONS (30 marks)

Choose the BEST option to answer the questions.

1. These are the objectives of learning histological techniques, EXCEPT

A. to operate the tissue processor, tissue embedding centre, rotary microtome and the floatation bath.

B. to study the characteristics of tissue microscopically.*

C. to stain sections with Haematoxylin and Eosin and other special stains.

D. to fix tissues in fixative solutions and decalcificate hard tissues.

2. These are the importances of specimen logging in histological techniques EXCEPT

A. to easily track the missing specimens during the processes

B. to obtain microscopically good slides at the end of the processes

C. to avoid processing wrong specimens from wrong patients

D. to claim the laboratory / hospital fees from patients during discharge*

3. Which of these terms refer to ‘bacterial attack’?

A. Purification

B. Putrefaction*

C. Autolysis--self-destruction

D. Cytolysis

4. Which of these acids is a weak acid that can be used in decalcification?

A. Nitric acid

B. Hydrochloric acid

C. Sulphuric acid

D. Acetic acid*

5. Which of these specimens need decalcification prior to fixation process?

A. Lungs

B. Kidneys

C. Skull*

D. Liver

6. Following are the aims of tissue processing EXCEPT

A. to embed tissues in solid medium.

B. to remove water so that paraffin wax can impregnate in the tissues.

C. to preserve tissue substances and proteins.*--fixation

D. to enable the knife to cut the section with little damage to knife or tissues.

7. Tissue processing steps involve in

A. hydration, clearing, infiltration and impregnation, and embedding.

B. hydration, cleaning, deparaffinization and embedding.

C. dehydration, clearing, infiltration and impregnation, and embedding.*

D. dehydration, clearing, deparaffinization and sectioning.

8. The following are properties of ethyl alcohol (ethanol) EXCEPT it is

A. flammable.*

B. has pleasant odour.

C. miscible with water.

D. brownish in colour.

9. Methanol properties are almost similar to ethanol EXCEPT it

A. is not miscible with water.*

B. has an unpleasant odour.

C. is non-flammable.

D. is yellowish in colour.

10. Clearing agent is removed from the tissue and replaced by wax in a process called

A. clearing.

B. infiltration and impregnation.*

C. dehydration.

D. embedding.

11. Too high temperature in infiltration and impregnation procedure will

A. over harden the tissues.*

B. molten the tissues.

C. increase the size of tissues.

D. enhance the bacterial growth within the tissues.

12. After embedding, the wax must be rapidly cooled to reduce the

A. wax crystal.*

B. wax ice.

C. wax salt.

D. wax iron.

13. Manual tissue processing is required in the following situations EXCEPT

A. for speed processing.

B. electrical power failure.*

C. during peak hours in lab.

D. machine breakdown.

14. “To expose suitable area of tissue for sectioning”. Choose an answer below that has the same meaning with that statement.

A. Embedding

B. Clearing

C. Fishing

D. Trimming*

15. The duration for routine automated tissue processing is

A. 10 hours.

B. 15 hours.

C. 20 hours.*

D. 25 hours.

16. Prolonged floating out of section on the water bath must be avoided as tissues may

A. expand.*

B. shrink.

C. flatten.

D. compress.

17. Picking up of sections from water bath is called

A. touching.

B. floating.

C. fishing.*

D. sliding.

18. What is the advantage when the waterbath has a black case?

A. The cost is much cheaper compared to clear case waterbath.

B. Sections floated in it will be more easily seen.*

C. It takes shorter time to warm to the desired temperature.

D. It can be used as an incubator as well.

19. These are the types of haematoxylin dye EXCEPT

A. iron haematoxylin.

B. phospomolybdic acid haematoxylin.

C. picric haematoxylin.*

D. lead haematoxylin.

20. Harris’s haematoxylin is the type of

A. haematoxylin without mordant.

B. alum haematoxylin.*

C. tungsten haematoxylin.

D. mercurial haematoxylin.

21. 0.5% ammonia in Haematoxylin and Eosin staining step is also called a ________________ solution.

A. greening

B. blacking

C. browning

D. bluing*

22. These are the types of eosin dye EXCEPT

A. eosin B.

B. eosin D.*

C. eosin Y.

D. ethyl eosin.

23. Which of these combinations between tissue structures and the H&E staining results is NOT TRUE?

A. Nuclei – blue black.

B. Cytoplasm – pink.

C. Muscle fibre – blue.*--pinky red

D. RBC – orange / red.

24. What is the most commonly used mounting media?

A. UHU glue.

B. Cellulose starch.

C. DPX resin.*

D. Lysin X.

25. The Congo Red stain is mainly used in the histological diagnosis of

A. dibetes mellitus.

B. tuberculosis.

C. Alzheimer disease.

D. amyloidosis.*

26. Which of these special stains is useful in the cases of mesotheliomas?

A. Von Kossa.

B. Oil Red O.

C. Masson Fontana.

D. Alcian Blue.*

27. Sudan Black stain is also known as

A. Oil red O stain.*

B. Gram stain.

C. Von Kossa stain.

D. Toluidine Blue stain.

28. Melanin and argentaffin molecules in tissues can be stained by using _______________ staining method.

A. Masson Trichrome

B. Gordon and Sweet’s

C. Von Kossa

D. Fontana-Masson*

29. Histology laboratory may receive the specimens in the form of

i. autopsies

ii. biopsies

iii. whole organs

iv. body fluids

A. i and ii

B. ii and iii

C. i, ii and iii*

D. all of the above

30. The aims of using vacuum in an automated tissue processor is to

i. remove air bubbles in the tissue.

ii. remove abnormal proteins in the tissue.

iii. remove clearing agents.

iv. increase the strength of tissue fibres

A. i and iii*

B. i and ii

C. i, ii, and iii

D. i, iii and iv

PART B – STRUCTURED QUESTIONS (40 marks)

Answer ALL questions.

Question 1

a. Give definition of fixative (2 marks)

b. Describe FOUR (4) advantages and disadvantages of 10% formol-saline fixative (8 marks)

Question 2

a. Define tissue processing (2 marks)

b. Explain the steps of embedding (6 marks)

c. Name two clearing agents other than xylene and two embedding media other than paraffin wax (2 marks)

Question 3

Describe FIVE (5) procedure outlines for paraffin sections staining and briefly explain each (10 marks)

Question 4

a. Name the preferred special stain to stain these components

i. Lipids

ii. Argentaffin granules

iii. Mucins

iv. Amyloids

v. Hemosiderin iron

vi. Melanin

vii. Calcium (7 marks)

b. List THREE (3) causes and remedies when sections roll into a coil (3 marks)

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