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Histo 2 > Exam 1 > Flashcards

Exam 1 Flashcards

1
Q

Basic Steps of Microtome Sectioning

A 
Embed tissue in parafin 
Face in block at 20 um
Chill/soften block on ice
Cut ribbons at 5 microns
Place on warm water bath 
Pick up on labeled slide
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2
Q

Types of Infectious Waste

A

Microbiological or culture material
Pathological Material
Blood
Sharp Objects

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3
Q

Types of Mechanical Hazards

A 
Blades
Needles
Scalpels
Razors
Glassware
Electrical Equipment
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4
Q

Types of Health Hazards

A

Infectious Disease
Carcinogens
Cryogenic Sprays
Toxins (Reproductive)

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5
Q

Types of Chemical Hazards

A 
Corrosives
Irritants
Sensitizers
Toxins
Carcinogens
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6
Q

Types of Fire Hazards

A

Alcohol

Hydrocarbons

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7
Q

Microtome vs. Cryostat: Temperature

A

Microtome: Room Temp
Cryostat: -20 degrees C

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8
Q

Microtome vs. Cryostat: Embedding Agent

A

Microtome: Parrafin
Cryostat: OCT

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9
Q

Microtome vs. Cryostat: Antiroll Plate (Y/N)

A

Microtome: No
Cryostat: Yes

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10
Q

Microtome vs. Cryostat: Water Bath

A

Microtome: Yes
Cryostat: No

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11
Q

Basic Steps of H&E

A 
Deparaffinization
Hydration
Nuclear Staining (Hematoxylin) 
Differentiation
Bluing
Counterstaining (Eosin)
Dehydration
Clearing 
(water rinsing steps in between)
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12
Q

What is the purpose of xylene in an H&E stain?

A

removes paraffin wax

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13
Q

How do you rehydrate the tissue for staining?

A

graded alcohols to water

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14
Q

What causes nuclear staining?

A

Hematoxylin

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15
Q

What causes differentiation in an H&E stain?

A

acid alcohol

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16
Q

What causes bluing in an H&E stain?

A

Ammonia Water (Alkaline water)

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17
Q

What is used for counterstaining in an H&E stain?

A

Eosin

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18
Q

What causes dehydration in an H&E stain?

A

Application of graded alcohol to 100% alcohol ( 95%, 95%, 100%, 100%)

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19
Q

How do you clear at the end of H&E staining?

A

Xylene

Transition from alcohol to non-aqueous reagents

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20
Q

Define: Regressive Staining

A

Tissue is overstained and then partially decolorized (differentiated) until the proper endpoint is reached
Faster and More convenient

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21
Q

Which staining type gives you a sharper degree of differentiation?

A

Regressive Staining

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22
Q

How is differentiation controlled in regressive staining?

A

By microscopic examination

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23
Q

Define: Progressive Staining

A

Tissue is stained for a predetermined time for adequate staining of the nuclei and leaves the background tissue relatively unstained

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24
Q

How does progressive staining work?

A

Once the dye is taken up by the tissues, it is not removed

The tissue is left in the dye solution until it retains the desires amount of coloration

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25
Q

How does differentiation work in progressive staining?

A

It relies on the selective affinity of dyes for different tissue elements

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26
Q

What are the components of Hematoxylin?

A 
Dye
Oxidizer
Mordant
Acidifier
Stabilizer
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27
Q

What does the dye in an H&E do?

A

it stains the different parts of the cells

28
Q

What does the oxidizer in an H&E do?

A

It converts hematoxylin to hematein

29
Q

What does the mordant in an H&E do?

A

Enhances binding of tissue to the slide

30
Q

What does the acidifer in an H&E do?

A

Changes pH

Helps with differentiation

31
Q

What does the stabilizer in an H&E do?

A

Decreases the rate of oxidizing

32
Q

What is the purpose of eosin?

A

It stains cytoplasmic (positive)

33
Q

What is the purpose of Hematoxylin?

A

It stains nuclear (negative)

Like DNA

34
Q

How do you remove eosin?

A

With alcohol

35
Q

What causes cell shrinkage?

A

overdehydrated tissue

36
Q

How do you recognize cell shrinkage?

A

Edges of the “holes” are not as jagged as torn tissue

37
Q

What causes faded/muddy nuclei?

A

Poor Fixation

38
Q

How do you recognize faded/muddy nuclei?

A

There will be no clear nuclei visible in the center of the tissue

39
Q

What causes moth-eaten holes?

A

dull knife or poorly processed tissue

40
Q

How do you recognize moth-eaten holes?

A

Holes look torn and are throughout the majority of the tissue

41
Q

How do tell the difference between cell shrinkage and moth-eaten?

A

Cell shrinkage: “holes” from overdehydrated tissue are uniform around the edges without torn appearance
Moth eaten: Holes look torn and are throughout the majority of the tissue

42
Q

How do you tell where a bubble is?

A

Bubbles under the tissue stain darker in the “bubble” region and can tear the tissue when they dry and pop - NOT FIXABLE
Mounting Bubbles do not damage the tissue, bubble appears unfocused - FIXABLE

43
Q

What causes wrinkles?

A

not properly stretching tissue on the water bath

44
Q

What causes folds?

A

picking-up tissue incorrectly from water bath onto slide or tissue not adhering properly to the slide and folding during staining

45
Q

What causes knife lines?

A

nicked/damaged blade

46
Q

How do you recognize Knife Lines?

A

The tissue looks torn in one distinct area and continues the length of the tissue

47
Q

What causes Venetian blinding chatter or washboarding?

A

loose or worn microtome parts (usually the blade is not locked)

48
Q

How do you recognize venetian blinding chatter/washboarding?

A

the light/dark/light/dark sections across the width of the tissue

49
Q

What causes parched earth in a tissue?

A

incomplete fixation

fixative moves from the outside of the tissue in - not enough time to penetrate the tissue or tissue is too dense

50
Q

How do you recognize parched earth?

A

the tissue is only damaged in the center

51
Q

What causes microchatter?

A

overdehydration (not fixable) or cutting too rapidly (fixable)

52
Q

How do you recognize microchatter?

A

tissue chatter is throughout the entire tissue

53
Q

What causes tearing?

A

overstretching on a water bath

knife lines

54
Q

What causes separation?

A

overstretching of tissue on water bath

incomplete fixation

55
Q

What causes Thick and Thin Sectioning?

A

Loose block or knife

Improperly embedded tissue

56
Q

You realized that the water solution you used to rinse your H&E slides was too alkaline. What would you expect to see with respect to your H&E staining when you look at your slides under the microscope?

A

Residual alkalinity can remove or damage tissue

57
Q

When observing your H&E stained slides under the microscope you notice blue-black “spots” throughout the entire slide. What is the most likely cause of this artifact?

A

The blue-black spots are caused by using hematoxylin that is expired or improperly stored.

58
Q

Microscopic analysis of H&E stained tissue shows brown nuclei. What is the most likely cause of this staining observation?

A

Brown nuclei could indicate that the hematoxylin used is breaking down or the sections aren’t sufficiently blued.

59
Q

You perform routine H&E staining on a set of slides. Upon microscopic examination you notice a number of unstained portions throughout some of the tissues. What is the most likely cause of this staining observation?

A

The most likely cause of this observation is from not fully removing the paraffin before staining.

60
Q

You are starting to feel comfortable working in the histology laboratory after starting your position three months ago. You decide to tweak the H&E staining protocol and start by increasing the time of the eosin stain from 5 minutes to 15 minutes. Your supervisor looks at your stained slides and gets upset telling you that there is a problem with the tissue slides you have generated. What is the most likely issue you will see when you look into the microscope?

A

Increasing the time the slides remain in the eosin can cause it to stain cytoplasmic structures too intensely making it hard to differentiate between things like red cells, collagen, and smooth muscle.

There wont be three shades of eosin

61
Q

Define: Cornflaking

A

Drying of slide before coverslipping

62
Q

What causes reddish brown nuclei?

A

Bluing Issue

63
Q

Eosin is __ charged and stains ___.

A

Eosin is negatively charged and stains positive

64
Q

Hematoxylin is ___ charged and stains __

A

Hematoxylin is positively charged and stains negative (DNA and Nucleic Acids)

65
Q

What is the differentiating agent do? What does it do?

A

Aci Alcohol

It removes excess hematoxylin

Histo 2 (4 decks)

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