Lesson 1: CYTO AND HISTOTECHNIQUES (ADVANCE CLASS)

Question 1

rooms in histopathology lab should have?

Answer 1

proper ventilation and good lighting

Question 2

Wax is heated at ___ in the embedding
machine

Answer 2

60C

Question 3

color of tissue cassette?
Skin

Answer 3

Gray

Question 4

color of tissue cassette?
Lymph nodes

Answer 4

pink

Question 5

Processes
that an automated machine can perform

Answer 5

FDCI (Fixation, Dehydration, Clearing, Infiltration)

Question 6

sections produced by a microtome

Answer 6

Ribbons

Question 7

color of tissue cassette?
-Liver/renal

Answer 7

Yellow

Question 8

color of tissue cassette?
Routine (e.g. placenta/uterus)

Answer 8

Green

Question 9

color of tissue cassette?
Bones

Answer 9

White

Question 10

Fresh tissue examination procedure

Answer 10

A. Teasing or Dissociation
B. Squash Preparation
C. Smear Preparation

Question 11

Smear Preparation procedure

Answer 11

➢ Streaking (zigzag manner)
➢ Spreading (spreading in a circular motion)
➢ Pull-apart
➢ Touch Preparation

Question 12

FIXATION
Tissue-to-Fixative Ratio–

Answer 12

1:10 or 1:20

Question 13

Accomplished by preserving and carefully processing
solid structures and tissues in the following order:

Answer 13

1. Fixation
2. Decalcification (OPTIONAL)
3. Dehydration
4. Clearing
5. Impregnation (INFILTRATION)
6. Embedding
7. Trimming
8. Microtomy
9. Staining
10. Mounting
11. Labeling

Question 14

Gas produced by oxidation of methanol

Answer 14

Formaldehyde

Question 15

Folmaldehyde 100% concentration?

Answer 15

gas form

Question 16

Folmaldehyde 37-40% concentration?

Answer 16

stock concentration (causes overhardening of the external surfaces of tissues)

Question 17

working solution; MOST COMMONLY USED IN PH
-Takes longer to prepare

Answer 17

10% Neutral Buffered Formalin

Question 18

10% Neutral Buffered Formalin
Fixation Time:

Answer 18

4-24 hours

Question 19

BEST GENERAL TISSUE FIXATIVE

-Best for iron-containing pigments and elastic fibers
which do not stain well after Susa, Zenker, or
Chromate fixation

Answer 19

10% Neutral Buffered Formalin

Question 20

Thickness of Specimen

○ Light Microscope:___
○ Electron Microscope: _____

Answer 20

2 cm2 x 0.4 cm2
1-2 mm2

Question 21

Temperature
○ Room Temperature ______: ROUTINE

Answer 21

(22C–28C)–40C

Question 22

factors affecting fixation

Answer 22

● Fixative of choice
● Time (20-30 minutes)
● Tissue-to-Fixative Ratio
● Penetration Rate (1 mL/hr)
● Thickness of Specimen
● Tissue Components
● pH
● Temperature
● Osmolality
● Agitation, Vacuum

Question 23

Temperature
○ Tissue Processors: ___

Answer 23

45C

Question 24

Temperature
○ Microwave: ___

Answer 24

65C

Question 25

Temperature ○ Tuberculosed Tissues: ____

Answer 25

100C

Question 26

Temperature ○ Electron Microscope: ____

Answer 26

0C–40C

Question 27

MOST COMMONLY USED ALCOHOL IN DEHYDRATION?

Answer 27

Ethanol

Question 28

Osmolality ○ MOST OPTIMAL: ______

Answer 28

400–450 mOsm (milliosmole)

Question 29

Tissue-to-Dehydrating Agent Ratio

Answer 29

1:10

Question 30

Temperature ○ Rapid Biopsy: ___

Answer 30

60C

Question 31

● Removal of fixative and water from the tissue ● Carried out by passing the tissues through a series of ascending grades of alcohol_________

Answer 31

Dehydration 70%, 90%, and absolute alcohol

Question 32

Decalcification done for how many hours?

Answer 32

48 hours

Question 33

Removal of calcium ions from a bone or calcified tissue through a histological process that makes them flexible and easier to cut

Answer 33

Decalcification

Question 34

most commonly employed medium in histopathology

Answer 34

Paraffin Wax

Question 35

Can cause damage to knife, poor trimming of tissues

Answer 35

INADEQUATELY DONE

Question 36

Tissue-to-Decalcifying Agent Ratio

Answer 36

1:120

Question 37

factors influencing the rate of decalcification

Answer 37

1. CONCENTRATION 2. FLUID ACCESS 3. SIZE AND CONSISTENCY 4. AGITATION 5. TEMPERATURE

Question 38

Completely transparent tissue indicates the _____ of the clearing process

Answer 38

TERMINAL POINT

Question 39

● AKA dealcoholization ● Process of removing the dehydrating agent from the tissue to facilitate impregnation of paraffin wax ● MAIN GOAL: Make tissues transparent/translucent

Answer 39

clearing

Question 40

FROZEN SECTIONS can use _____ and ____ as dehydrating agent

Answer 40

glycerin, gum syrup

Question 41

xylene 1 and 2 time interval?

Answer 41

20 minutes

Question 42

xylene 3 time interval

Answer 42

45 minutes

Question 43

○ Commonly employed clearing agent; cheapest ○ Clear fluid as well as a cheap and excellent lipid solvent ○ Becomes milky if tissue is not properly dehydrated

Answer 43

Xylene

Question 44

Infiltration and embedding Duration per Section:

Answer 44

1 hour–1 hour–3 hours

Question 45

-Impregnate in a medium that could support the tissue -For easier handling of tissue

Answer 45

infiltration and embedding

Question 46

Infiltration and embedding ○ Wax/Paraffin Oven: ____ than melting point ○ Wax can only be reused _____

Answer 46

>2C–5C twice

Question 47

INFILTRATION Procedure Immerse the tissue samples in 3 stages of paraffin wax following the given time intervals respectively: ● 1st change paraffin wax bath: _____ ● 2nd change paraffin wax bath: ______ ● 3rd change paraffin wax bath: _____

Answer 47

30 minutes 30 minutes 45 minutes

Question 48

Paraffin wax may be used only twice - Removed by heating the wax to _____

Answer 48

100-105C

Question 49

Temperature of embedding paraffin wax?

Answer 49

2-4C above its melting point

Question 50

● Arrangement of the tissue in a precise position in the mold during embedding ● MOST CRUCIAL STEP DURING EMBEDDING

Answer 50

Orientation

Question 51

OVERHEATING OF WAX cause

Answer 51

○ Hardening and distortion of the tissue ○ Breakdown of the paraffinic additive (e..g resin polymer)

Question 52

usually form ribbons due to slight heat generated between the block and the knife edge during the process of cutting

Answer 52

SECTIONS

Question 53

Usual thickness of tissue in sectioning

Answer 53

10 microns

Question 54

AFTER MICROTOMY Heat the slide gently on the slide warmer (_____) until the ribbons spread and flatten ou

Answer 54

40-45C

Question 55

Floatation Bath:____ lower than temperature of paraffin wax

Answer 55

5-10C

Question 56

Spring-balance or pawl is brought into contact with the block, and turns a ratchet feed wheel connected to a micrometer screw, which is in turn rotated, moving the tissue block at a predetermined distance towards the knife for cutting sections at uniform thickness

Answer 56

Microtome

Question 57

used to clean the knife

Answer 57

Xylene/Wax Paper/Brush

Question 58

Process of applying des on the section to see and study the architectural pattern of the tissue and physical characteristics of the cells

Answer 58

staining

Question 59

basic, attracted to acidic part (nucleus) of the cell -Stains purple-blue

Answer 59

Hematoxylin Stain

Question 60

acidic, attracted to basic part (cytoplasm) of the cell -Stains red-orange

Answer 60

Eosin Stain

Question 61

Rehydration: descending grades of alcohol________

Answer 61

(100%–90%–70%)

Question 62

Routine H&E Staining procedure

Answer 62

STEP 1 Deparaffinization STEP 2 Rehydration: Graded alcohol STEP 3 Nuclear stain: Haematoxylin STEP 4 Differentiation STEP 5 Bluing STEP 6 Counterstain: Eosin STEP 7 Dehydration STEP 8 Clearing and Mounting

Question 63

In Routine H&E Staining procedure what is the nuclear stain?

Answer 63

Haematoxylin

Question 64

In Routine H&E Staining procedure what is the counterstain?

Answer 64

eosin

Question 65

Process of removing excess hydrogen ion from the stain

Answer 65

BLUING

Question 66

Mounting Protective layer:

Answer 66

between 1.0 and 2.5 mm thick

Question 67

Process of placing the coverslip on the slide with the use of a mounting media

Answer 67

Mounting