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HistoPathology > [M] Week 8: Gross Room / Surgical Cut- up - Part 1 > Flashcards

[M] Week 8: Gross Room / Surgical Cut- up - Part 1 Flashcards

1
Q

Tissues from the body taken for diagnosis of disease processes must be processed in the histopathology laboratory to produce ____ ____ that are viewed under the microscope by pathologists

A

Microscopic slides

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2
Q

The persons who do the tissue processing and make the glass microscopic slides are

A

histotechnologists

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3
Q

In the laboratory safety precautions must observed at all times because we are dealing with a lot of hazards like?

A
  • Biological Hazard
  • Chemicals
  • Radiation

Safety first and last!

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4
Q
  • The interface between hospital staff (or other visitors) and the pathology personnel (histotechnologists and other clerks that may receive the specimen)
  • To receive samples safely and securely
A

Specimen Reception

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5
Q

SPECIMEN HANDLING AND IDENTIFICATION

True/False
1. Specimen labeled properly attached to the cap of the container
2. Request form is needed

A
  1. False (should be in the body)
  2. True
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6
Q

SPECIMEN HANDLING AND IDENTIFICATION

What are included in the request form?

A
  • patient information
  • medical history
  • description of the site of origin
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7
Q

SPECIMEN HANDLING AND IDENTIFICATION

surgical pathology request form, contains what information needed?

A
  • patient info
  • clinical diagnosis
  • specimen site or laterality
  • operation or procedure done
  • examinations requested (histopathology, cytology, cell block for safekeeping)
  • pertinent history or diagnostics done on the patient
  • The requesting physician.

This information is filled out by the operating room personnel (doctors, residents, nurses).

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8
Q

SPECIMEN HANDLING AND IDENTIFICATION

In the laboratory, we receive the specimen so take note of
the ____ and ____ received and the person who received the specimen, then we will assign the ____

A
  1. date and time
  2. accession number
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9
Q

SPECIMEN ACCESSIONING

  • A complex number or alpha numeric number that will identify each specimen for each patient
  • Not universal for all institutions. O’thers may have a different way of assigning.
A

Accession Number

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10
Q

SPECIMEN ACCESSIONING

What is the format of accession number?

A

Procedure - Year - Chronology when spx is received

S for surgical
P for Pap’s smear for cytology
CB for cell block

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11
Q

After receiving a specimen and labelling it, what will you do next?

A

check if there is a fixative (such as formalin) in the container.

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12
Q

After fixing for several hours, the specimen will be?

A

grossed

like yung EWWW

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13
Q

specimen dissection area, should have?

A
  • Good lighting
  • Good ventilation
  • Non-absorbent wipe-clean surfaces
  • Appropriate protective clothing for the laboratory personnel (fluid resistant laboratory gown and masks for protection from fumes)
  • Gloves
  • Cutting tools
  • Other equipment (photography, tissue macerators, bone saw, trash bin)
  • Integrated dissection desks (Or a kitchen-like tabletop with tiles and a sink )
  • Enclosed fluid/fixative feeds
  • Laminar down-draft ventilation (Alternative: Exhaust fan)
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14
Q

Tissues are examined by a

A

pathologist, pathology assistant, or pathology resident

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15
Q

For gross examination, these are the things we have to take note of, what are those?

A
  • Describing the specimen
  • Placing all or just a part of the specimen in a tissue cassette
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16
Q

GROSS EXAMINATION

Number of tissue/s present in the specimen container, Shape, Color, Consistency, Size of the specimen (3-dimensional: LxWxH in cm)

A

Describing the specimen

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17
Q

GROSS EXAMINATION

Tissue inside the cassette should be dissected to?

A

3-4mm in thickness

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18
Q

gross only examination is done with what specimens?

A
  • Non-tissue
  • Tissues unlikely to require adetailed diagnosis and may be examined grossly
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19
Q

When a malignancy is suspected, then the specimen is covered with ink – to mark the margins of the specimen

A

Inking the Margins

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20
Q

INKING THE MARGINS

When sections are made and processed, the ink will mark
the ____ ____ on the slide

A

actual margin

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21
Q

INKING THE MARGINS

TOF

It is also used to see if the margin is free of malignant cells and also how far the malignant cells are from the margin.

A

True

The farther the malignant cells are from the margin, the safer.

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22
Q

TISSUE PROCESSING

After the removal of a tissue sample from the patient, a series of processes must take place to ensure the final microscopic slides are of?

A

diagnostic quality

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23
Q

TISSUE PROCESSING

TOF
Producing quality slides for diagnosis is an accident;

A

F

requires skills that are developed through continued practice
and experience.

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24
Q

TISSUE PROCESSING

Tissue processing aims to?

A
  • Permanently preserve the tissue
  • Stain tissue for demonstration of specific structures
  • Mount tissue on glass slides with coverslips for permanent keeping
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25
Q

TISSUE PROCESSING

Tissue processing is designed to remove all ____ ____ from the tissue, replacing it with a support medium that provides sufficient rigidity to enable sectioning of the tissue
without damage or distortion

A

Extractable Water

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26
Q

Give men the tissue processing steps

A
  1. Fixation (most critical step)
  2. Dehydration
  3. Clearing (De-alcoholization)
  4. Impregnation
  5. Embedding
  6. Trimming
  7. Section-cutting
  8. Staining
  9. Mounting
  10. Labeling
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27
Q

What is the usual or routine fixative used?

A

Formaline or Formaldehyde

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28
Q

For dehydration step, what are the routinely used dehydrating agents?

A

increasing grades of alcohol

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29
Q
  • In this step, we remove the alcohol added during dehydration.
  • After dehydration, the tissue will become transparent or clear. Hence, the other name for this step is clearing.
A

De-alcoholization

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30
Q

Since we removed all extractable water: holes, gaps, or spaces will be left in the tissue, making it difficult to cut through the tissue into thin sections. It has to have a support medium obtained by filling up the spaces. It is called?

A

impregnation or infiltration

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31
Q

For impregnation or Infiltration, what is routinely used?

A

Melted paraffin

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32
Q

The tissue that was impregnated will be placed in a mold, what will be added?

A

embedding medium

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33
Q

We are now ready to cut into thin sections. Sometimes tissue blocks will be too big for the block holder, so the excess needs to be removed by?

A

trimming

do the coarse cutting cutting the tissue by 10 to 15 microns thickness

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34
Q

When tissue is already exposed, we will adjust the thickness to 4 to 6 microns to cut into tissue or?

A

serial sections

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35
Q

Then, sections will be placed on a glass slide to be stained using routinely used

A

H&E

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36
Q

After staining what will be done?

A

Mounted, using mounting medium

then label after (last step)

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37
Q
A
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38
Q

For hard tissues (ex. bones, calcified tissues) what extra step are used?

A

decalcification (after fixation & before dehydration)

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39
Q

DECALCIFICATION

Hard tissues contain? which make them hard

A

calcium salts or lime salts

Removing these salts will make the tissue softer and easier to cut into thin sections

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40
Q

routinely used decalcifying agent

A

10% nitric acid

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40
Q

yoooo

A

study ng tissue processing schedule

okay?

=

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41
Q

Tissue cassettes are manually transferred from one reagent to another

A

Manual tissue processing

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42
Q
  • Time is usually set with an alarm clock, cassettes are transferred when the time ends to the next reagent.
  • This is carried out at room temperature
A

Manual tissue processing

43
Q

automatic tissue processor, temperature is also employed.

This model contains reagents in different containers

44
Q

AUTOMATIC TISSUE PROCESSOR “TECHNICON”

used to hold the tissue cassettes

A

Tissue Basket

45
Q

AUTOMATIC TISSUE PROCESSOR “TECHNICON”

Time is set digitally at the ____ ____ of the Technicon, inputted through the digital pads, the timing for each step of tissue processing from when it starts to when it ends.

A

lower portion

46
Q

AUTOMATIC TISSUE PROCESSOR “TECHNICON”

the tissue processing up to impregnation will take?

A

6 to 8 hours

47
Q

AUTOMATIC TISSUE PROCESSOR “TECHNICON”

For this model, tissue cassettes are placed in a ____ and instead of the basket moving from one container to another, the reagent will be flooded in that area.

48
Q

AUTOMATIC TISSUE PROCESSOR “TECHNICON”

The baskets for both models employ agitation, by ____ ____ , and rotation, by ____ ____. These oscillations will aid in tissue processing, as well as the heat they create.

A
  1. Vertical Oscillation
  2. Rotary oscillation
49
Q

AUTOMATIC TISSUE PROCESSOR “TECHNICON”

Tissue cassettes contained in baskets through a series of reagents

Vertical oscillation or the mechanical ____ and ____ of the tissue and rotary oscillation provide the ____ needed

A
  1. Raising and lowering
  2. Agitation
50
Q
  • The tissue will be placed in molds that can be metal, plastic or disposable mold like the paper boat.
  • After the paraffin solidifies, a paraffin or tissue block will be produced, and later placed in a microtome.
51
Q

Section-Cutting

A microtome has different types, what are routinely used in the laboratory?

A

Minot or rotary microtome

It can be manually, mechanically, or automatically driven.

52
Q

Section-Cutting

An example of a non-automatic one. Regardless of whether it is manual or automatic, they all have these basic parts, what are those?

A
  • Block holder
  • Blade holder
  • Wheel (moves the block holder)
  • Knob (sets the thickness)
53
Q

Section-cutting

has a thickness of 4 to 6 microns which is later transferred to a water bath.

A

Tissue ribbon

53
Q

Floatation

The water bath is set at 6-10 degrees lower than the melting point of paraffin.

At this point, the paraffin should not melt, it should just be flattened to remove the wrinkles and will be separated into individual sections.

A

Floatation

54
Q

Sections are retrieved by ?

A

Fishing out
in a near-vertical position close
to the section it will attach to.

55
Q

This is used to melt the paraffin, making the tissue adhere more to the slide, like heat fixation.

A

Paraffin oven

56
Q

What is used for the staining of the slides?

A

hematoxylin and eosin (H&E)

57
Q

after the staining, what will be done?

58
Q

describe the mounting medium

A

Syrupy liquia covered with a coverslip

yum

59
Q

How much mounting medium is dispensed?

A

1 to 2 drops, enough to cover the entire tissue

60
Q

Mounting

TOF

After it solidifies or dries up, the coverslip will be permanently
placed. Sometimes, we place something to seal around the
coverslip to prevent the mounting medium from coming out.

61
Q

The slides are labeled according to?

A

Types of specimen and staining
But in actual practice, the labels include the accession number and the institution or hospital.

No need to write the name of
the patient nor the type of
specimen, only the institution or
hospital and the accession number.

62
Q

The size & type of specimen in the tissue cassette determine the need for complete fixation and processing, what is the thickness required?

A

Only 3-4mm thickness should be placed in the tissue cassette

Do not overfill the cassette so that it will not impede the reagents when we do the processing

63
Q
  • The first and most critical step of tissue processing in histotechnology
  • Involves fixing or preserving fresh tissue for examination
  • The entire process of tissue processing will depend on
    this step.
A

Fixation

If the tissue is fixed incompletely, overfixed, or the wrong fixative is used, it will cause autolysis, damaging the tissue.

64
Q

What is the primary aim of fixation?

A

To preserve the morphological and chemical integrity of the cell as life-like manner as possible

65
Q

What is the secondary goal of fixation?

A

To harden and protect the tissue from the trauma of further handling, so that it is easier to cut during gross examination

66
Q

What are the two mechanism involved in fixation?

A
  • Additive Fixation
  • Non-additive Fixation
66
Q

Enumerate why fixation is performed

A
  • To preserve the tissue
  • To prevent the breakdown of cellular elements
  • To coagulate or precipitate protoplasmic substances that may leak out during subsequent steps of tissue processing
  • As soon as possible after removal of the tissues (in the case of surgical pathology) or soon after death (with autopsy) to prevent autolysis
67
Q
  • The chemical constituent of the fixative is taken in and becomes part of the tissue by forming crosslinks or molecular complexes and giving stability to the protein
  • Ex. formalin, mercury, and osmium tetroxide
A

Additive Fixation

68
Q
  • The fixing agent is not incorporated into the tissue, but alters the tissue composition and stabilizes the tissue by removing the bound water attached to H-bonds of certain groups within the protein molecule
  • Ex. alcohol fixatives
A

Non-additive Fixation

69
Q

What are the Main factors Involved in fixation

A
  • Hydrogen Concentration
  • Temperature
  • Thickness of Section
  • Osmolality
  • Concentration
  • Duration of Fixation
70
Q

MAIN FACTORS INVOLVED IN FIXATION

pH 6 to 8

A

Hydrogen Ion Concentration

71
Q

MAIN FACTORS INVOLVED IN FIXATION

Provide the temperature needed

  • Tradtional Temp
  • Tissue processors
  • Electron microscopy and Histochemistry
A
  • RT
  • 40 degree C
  • 0-4 degree C
72
Q

MAIN FACTORS INVOLVED IN FIXATION

Small or Thin
1. 1 to 2 mm for electron miscroscopy
2. 2cm for light microscopy
3. No more than 0.4cm for light microscopy

A
  1. Small
  2. Small
  3. Thin
73
Q

MAIN FACTORS INVOLVED IN FIXATION

What measurement of osmolality will give the best result?

A
  • Slightly hypertonic solution (400 to 450 mOsm)
  • Isotonic solutions (340 mOsm)
74
Q

MAIN FACTORS INVOLVED IN FIXATION

What are the percentage used before the fixative

A
  • 10% formaldehyde
  • 3% glutaraldehyde
75
Q

MAIN FACTORS INVOLVED IN FIXATION

what is the duration of fixation?

A

2 to 6 hours
if prolonged causes shrinkage and hardening of tissue

76
Q

What are the practical consideration of Fixation

A
  1. Speed
  2. Penetration
  3. Volume
  4. Duration of fixation
77
Q

PRACTICAL CONSIDERATION OF FIXATION

  • Specimen should be placed in fixative as soon as it is removed from the body
  • Delaying fixation can cause autolysis
78
Q

PRACTICAL CONSIDERATION OF FIXATION

  • approximately 1mm/hr & slows down as it goes deeper
  • Cut large or solid organs into sections for the fixative to reach the deeper portions.
  • For hollow organs that tend to float, a gauze is soaked in fixative to prevent the specimen from floating.
A

Penetration

79
Q

PRACTICAL CONSIDERATION OF FIXATION

  • 10-25x the volume of tissue to be fixed
  • The tissue should be completely submerged in fixative
  • The size of the container is crucial. (Large specimens should not be put into smaller containers so that they can be filled with enough fixatives to soak them)
80
Q

What are the factors influencing the rate of processing?

A
  1. Agitation
  2. Heat
  3. Viscosity
  4. Vacuum
81
Q

FACTORS INFLUENCING THE RATE OF PROCESSING

  • The rate of exchange is dependent upon the exposed surface of the tissue that is in contact with the processing reagent
  • In automated processors, like Technicon, vertical or rotary oscillation or pressurized removal and replacement of fluids at timed intervals are employed,
82
Q

FACTORS INFLUENCING THE RATE OF PROCESSING

  • Increases the rate of penetration & fluid exchange
  • It must be used sparingly or with caution to reduce the possibility of shrinkage, hardening, and brittleness of the tissue
  • Temperatures limited to 45˚C can be used effectively; Higher temperature may be deleterious to tissues especially for immunohistochemical staining
83
Q

FACTORS INFLUENCING THE RATE OF PROCESSING

Is the property of resistance to the flow of a fluid

84
Q

FACTORS INFLUENCING THE RATE OF PROCESSING

The smaller the size of the molecules in the solution, the faster the rate of fluid penetration

A

Low viscosity

85
Q

FACTORS INFLUENCING THE RATE OF PROCESSING

If the molecular size is larger, the rate of exchange is slower

A

High viscosity

Smaller is faster; Larger is slower

86
Q

FACTORS INFLUENCING THE RATE OF PROCESSING

  • Using reduced pressure to increase the rate of infiltration
  • If used on the automated processor, it should not exceed 15 inches of Hg (mercury) to prevent damage and deterioration to the tissue
87
Q

EFFECTS OF FIXATIVES

True/False

  1. Soften hard & friable tissues and make handling and cutting sections easier
  2. Make cells resistant to damage & distortion
  3. Inhibit virus decomposition
A
  1. False (Harden soft)
  2. True
  3. False (bacterial decomposition)
88
Q

EFFECTS OF FIXATIVES

True/False

  1. Increase optical differentiation of cells and tissue components
  2. Act as mordants or accentuators to promote and hasten staining or inhibit certain dyes in favor of another
  3. Reduce risk of infection during handling and actual processing of tissues
A
  1. True
  2. True
  3. True
89
Q

CHARACTERISTICS OF A GOOD FIXATIVE

just familliarize

A
  • It must be cheap
  • It must be stable.
  • It must be safe to handle.
  • It must kill the cell quickly thereby producing minimum distortion of cell constituents.
  • It must inhibit bacterial decomposition and autolysis.
  • It must produce minimum shrinkage of tissue.
  • It must permit rapid and even penetration of tissues.
  • It must harden tissues thereby making the cutting of sections easier.
  • It must be isotonic, causing minimal physical and chemical alteration of the cells and their constituents.
  • It must make cellular components insoluble to hypotonic solutions and render them insensitive to subsequent processing.
  • It must permit the subsequent application of many staining procedures to facilitate easier and more profitable examination.
90
Q

What are the types of fixatives according to composition

A
  • Simple Fixatives
  • Compound Fixative
91
Q

TYPES OF FIXATIVES

  • made up of only one component substance
  • Aldehydes: formaldehyde, glutaraldehyde
  • Metallic fixatives: mercuric chloride, chromate fixatives, picric acid, acetic acid, acetone, alcohol, osmium tetroxide
A

Simple Fixative

92
Q

TYPES OF FIXATIVES

made up of two or more fixatives which have been added together to obtain the optimal combined effect of their individual actions upon the cells and tissue constituents

A

Compound Fixatives

93
Q

What are the types of fixatives according to action

A
  • Microanatomical Fixatives
  • Cytological Fixatives (Nuclear Fixatives, Cytoplasmic Fixatives)
  • Histochemical Fixatives
94
Q

Fixative according to action

  • Permit the general microscopic study of tissue structures without altering the structural pattern and normal intercellular relationship of the tissues in question
  • 10% Formal saline, 10% Neutral buffered formalin, Heidenhain’s Susa, Formal sublimate (formol corrosive), Zenker’s solution, Zenker-formal (Kelly’s solution), Bouin’s
    solution, Brasil’s solution
A

Microanotmical Fixative

95
Q

Fixative according to action

  • preserve specific parts and particular microscopic elements of the cell itself: nuclear or cytoplasmic components
A

CYTOLOGICAL FIXATIVES

96
Q

Fixative according to action

Cytological Fixative

  • preserve the nuclear structures (e.g., chromosomes)
  • usually contain glacial acetic acid
  • Flemming’s fluid w/ acetic acid, Carnoy’s fluid, Bouin’s fluid, Newcomer’s fluid, Heidenhain’s Susa
A

Nuclear Fixatives

97
Q

Fixative according to action

Cytological Fixative

  • preserve cytoplasmic structures
  • Flemming’s fluid without acetic acid, Kelly’s fluid, Formalin with “post-chroming”, Regaud’s fluid (Muller’s fluid), Orth’s fluid
A

Cytoplasmic Fixative

98
Q

Fixative according to action

  • preserve the chemical constituents of cells and tissues
  • 10% Formal saline, Absolute ethyl alcohol, Acetone, Newcomer’s fluid
A

Histochemical Fixatives

99
Q

Fixative according to action

what are the other fixative under histochemical fixatives?

A
  • Lipid Fixation
  • Carbohydrate Fixation
  • Protein Fixation
  • Glycogen Fixation
100
Q

HISTOCHEMICAL FIXATIVES

Cryostat or frozen section followed by general lipid stains

Mercuric chloride & Potassium
dichromate

A

Lipid Fixation

101
Q

HISTOCHEMICAL FIXATIVES

Carbohydrate fixation

A

Alcohol fixatives

102
Q

HISTOCHEMICAL FIXATIVES

Neutral buffered formalin, formol saline or formaldehyde

A

Protein fixation

103
Q

HISTOCHEMICAL FIXATIVES

Alcohol-based, such as Rossman’s fluid or cold absolute alcohol

A

Glycogen fixation

HistoPathology (14 decks)

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