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HISTOPATHOLOGY (TopnotcherQT) > FIXATION > Flashcards

FIXATION Flashcards

1
Q

Steps (in order) in tissue processing

A
  1. Fixation
  2. Decalcification
  3. Dehydration
  4. Clearing
  5. Infiltration or impregnation
  6. Embedding
  7. Trimming
  8. Sectioning or Cutting
  9. Staining
  10. Mounting
  11. Labeling

MNEMONIC:
Fuck DDis CIET Sige Share Mo Lang

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2
Q

Process of identifying the specimen WITHOUT WRITING THE PATIENT’S NAME on the specimen tag

A

NUMBERING

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3
Q

In numbering:

S is for ________
A is for ________
C is for _______

A

S is for SURGERY
A is for AUTOPSY
C is for CYTOLOGY

NOTE:

YEAR is indicated usually in 2 digits
ACCESSION NUMBER
PENCIL is utilized to label

Example: S23-143
S - Surgery
23 - Year sample received
143 - Accession number

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4
Q

FIRST and MOST CRITICAL STEP in routine tissue processing; The process of stopping all cellular activities so that cells can be viewed under the microscope

A

FIXATION

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5
Q

Most important reaction in maintaining the morphology of the tissue in fixation

A

Stabilization of proteins

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6
Q

PRIMARY AIM OF FIXATION:

A

Preserve the MORPHOLOGICAL AND CHEMICAL INTEGRITY of the cell in as life-like manner as possible

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7
Q

SECONDARY AIM OF FIXATION:

A

HARDEN AND PROTECT the tissue from trauma of further handling, so that it is easier to cut during gross examination

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8
Q

Main factors involved in fixation:

A
  1. Speed
  2. Rate of penetration
  3. pH
  4. Temperature
  5. Thickness of section
  6. Osmolality
  7. Concentration
  8. Amount of fixative
  9. Duration of fixation
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9
Q

Speed for fixation:

A

IMMEDIATELY fix after removal

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10
Q

Rate of penetration of fixation:

A

1 mm/hr for formalin from periphery to interior

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11
Q

pH of fixatives should be:

A

NEUTRAL (between 6 and 8)

BEST pH: 7 (formalin pH)

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12
Q

Temperature for fixation

Routine tissue processing:
For urgent biopsies/rapid fixation:
Electron microscope (EM):
Automatic tissue processor:

A

Routine tissue processing: ROOM TEMPERATURE
For urgent biopsies/rapid fixation: 60C
Electron microscope (EM): 0 to 4C
Automatic tissue processor: 40 to 42C

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13
Q

Thickness of section

Recommended size of the tissue:
Electron microscope (EM):
Light microscopy:

A

Thickness of section

Recommended size of the tissue:
- 2cm^2 and no more than 4mm thick

Electron microscope (EM): 1-2mm thick

Light microscopy: 4mm thick

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14
Q

Osmolality in fixation:

A

SLIGHTLY HYPERTONIC SOLUTIONS usually at 400-450 mOsm

<400 = SWELL
>450 = SHRINK

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15
Q

Concentration of fixatives

Formalin:
Glutaraldehyde:

A

Concentration of fixatives

Formalin: 10% working solution
Glutaraldehyde: 3% normal solution; for EM: 0.25% solution

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16
Q

Amount of fixative

Routine tissue processing:
Osmium tetroxide:
Electron microscope (EM):
Museum preparation:

A

Amount of fixative

Routine tissue processing: 20 TIMES the volume of the tissue

Osmium tetroxide: 5 to 10 TIMES the volume of the tissue

Electron microscope (EM): 20 TIMES the volume of the tissue

Museum preparation: <50 TIMES the volume of the tissue

NOTE:
RATIO OF TISSUE TO FIXATIVE = 1:20
RATIO OF FIXATIVE TO TISSUE = 20:1
**Traditionally 10-20 times the tissue

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17
Q

Duration of fixation:

A

Adequate fixation is 4 to 6 hours

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18
Q

Prolonged fixation can lead to:

A

SHRINKAGE
TOO MUCH HARDENING
INHIBITS CHEMICAL REACTIONS

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19
Q

Factors that HASTENS/ENHANCE fixation:

A
  1. Thin tissue sections
  2. Agitation
  3. Moderated heat (room temperature)
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20
Q

Factors that RETARDS/SLOWS fixation:

A
  1. Thick tissue sections
  2. Presence of FATS, MUCUS, and BLOOD
  3. Cold temperature
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21
Q

Remedies of there are presence of fats mucus and blood in tissue sections:

A

FATS - cut the tissue into smaller pieces
MUCUS and BLOOD - wash with NSS

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22
Q

Characteristics of GOOD FIXATIVE

A
  1. Cheap, stable and safe to handle
  2. Kills cells quickly; must inhibit bacterial decomposition and autolysis
  3. Make tissue resistant to damage and distortion
  4. Produce minimum shrinkage of tissue
  5. Rapid and even penetration of tissues
  6. Harden the tissue
  7. Must make cellular components INsoluble to hypotonic solutions and render them INSENSITIVE to subsequent processing (RECALL)
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23
Q

TWO (2) mechanisms of fixation:

A
  1. Additive fixation
  2. Non-additive fixation
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24
Q

Chemical constituent of the fixative is TAKEN IN and becomes a part of tissue

A

ADDITIVE FIXATION

25
Q

Examples of fixatives in additive fixation:

A
  1. Formaldehyde fixatives
  2. Mercuric chloride fixatives
  3. Osmium tetroxide fixatives
26
Q

Fixing agent is NOT TAKEN into the tissue but it alters the tissue composition thus, stabilizing the tissue which makes them unstainable for bacterial decomposition

A

NON-ADDITIVE FIXATION

27
Q

Example of a Non-additive fixative:

A

Alcohol fixatives

28
Q

FIXATIVES according to composition:

A
  1. Simple fixatives
  2. Compound fixatives
29
Q

Fixative that is made up of ONLY ONE component of substance

A

SIMPLE FIXATIVES

30
Q

Examples of SIMPLE FIXATIVES:

A
  1. Aldehyde fixatives
  2. Metallic fixatives
  3. Picric acid fixatives
  4. Alcohol fixatives
  5. Osmium tetroxide (Osmic acid)

mnemonic: AMPAO

31
Q

Fixatives under Aldehyde fixatives:

A
  1. FORMALDEHYDE
    - 10% formol saline
    - 10% Neutral buffered formalin or phosphate buffered formalin (pH7)
    - Formol-corrosive (formol sublimate)
    - Alcohol-formalin (Gendre’s fixative)
  2. GLUTARALDEHYDE

mnemonic:
Feeling Good si Aldehyde

32
Q

Fixatives under Metallic fixatives:

A
  1. MERCURIC CHLORIDE
  2. CHROMATE FIXATIVES
  3. LEAD FIXATIVES
    mnemonic = madaming METALS sa Malaysia, China, at Laos
  4. MERCURIC CHLORIDE
    - B-5 fixative
    - Heidenhain’s Susa solution
    - Zenker’s fluid
    - Zenker-formol (Hely’s fluid)
    mnemonic = ang mga tao sa Malysia ay laging BHZZe (busy)
  5. CHROMATE FIXATIVES
    - Chromic acid
    - Regaud’s fluid
    - Orth’s fluid
    - Potassium dichromate
    mnemonic = ang mga tao sa China ay laging naka-CROP top
  6. LEAD FIXATIVES
    mnemonic = masarap mag-isa sa Laos
33
Q

Fixatives under Picric acid:

A
  1. Bouin’s solution
  2. Brasil’s alcoholic picroformol

mnemonic: BaBy ni Picric acid

34
Q

Fixatives under alcohol fixatives:

A
  1. Isopropyl alcohol 95%
  2. Carnoy’s fluid
  3. Ethyl alcohol 70-100%
  4. Methyl alcohol 100%
  5. Newcomer’s fluid

mnemonic: ICE MuNa bago ALCOHOL

35
Q

Fixatives under Osmium tetroxide (Osmium acid)

A
  1. Flemming’s solution
  2. Flemming’s solution without acetic acid

Feel na Feel ni Osmium tetroxide

REMEMBER: Glacial acetic acid - goods na goods sa nucleus, bad na bad sa cytoplasm

**Nuclear fixatives - WITH Glacial acetic acid
**Cytoplasm fixatives - WITHOUT Glacial acetic acid

36
Q

REMEMBER: Glacial acetic acid - goods na goods sa nucleus, bad na bad sa cytoplasm

**Nuclear fixatives - WITH Glacial acetic acid
**Cytoplasm fixatives - WITHOUT Glacial acetic acid

37
Q

Made up of TWO or MORE fixatives which have been added together

A

COMPOUND FIXATIVES

38
Q

Bouin’s solution is a compound fixative that contains:

A
  • Formaldehyde
  • Picric acid
  • Glacial acetic acid
39
Q

Carnoy’s fluid is a compound fixative that contains:

A
  • Glacial acetic acid
  • Absolute alcohol
  • Chloroform
40
Q

FIXATIVES according to Action

A
  1. Microanatomical fixatives
  2. Histochemical fixatives
  3. Cytological fixatives
41
Q

Permit the GENERAL microscopic study of tissue structures

A

MICROANATOMICAL FIXATIVES

42
Q

Microanatomical fixatives should never contain _____________ because it inhibits hematoxylin

A

Microanatomical fixatives should NEVER CONTAIN OSMIUM TETROXIDE (OSMIC ACID) because it inhibits hematoxylin

43
Q

Examples of Microanatomical fixatives:

A

10% Formol Saline
10% Neutral Buffered Formalin
Heidenhain’s Susa
Formol sublimate (Formol corrosive)
Zenker’s solution
Zenker-Formol (Helly’s)
Bouin’s solution
Brasil’s solution

Mnemonic:
1010HF
ZZBB

44
Q

Preserve the CHEMICAL constituents of cells and tissues:

A

HISTOCHEMICAL FIXATIVES

45
Q

Examples of Histochemical fixatives:

A

10% Formol Saline
Absolute ethyl Alcohol
Acetone
Newcomer’s

mnemonic: FAAN

46
Q

Preserves SPECIFIC parts and particular microscopic element of the cell

A

CYTOLOGICAL FIXATIVES

47
Q

Examples of Cytological fixatives

A

A. NUCLEAR FIXATIVES
B. CYTOPLASMIC FIXATIVES

48
Q

Nuclear fixatives usually contains __________________ due to its affinity for nuclear chromatin

A

CONTAINS GLACIAL ACETIC ACID

REMEMBER: Glacial acetic acid - goods na goods sa nucleus, bad na bad sa cytoplasm

**Nuclear fixatives - WITH Glacial acetic acid
**Cytoplasm fixatives - WITHOUT Glacial acetic acid

49
Q

pH of Nuclear fixatives should have:

A

pH of 4.6 or less (<4.6)

50
Q

Examples of Nuclear fixatives:

A

Bouin’s solution
Flemming’s with glacial acetic acid
Carnoy’s
Newcomer’s
Heidenhain’s Susa

mnemonic:
BE(F)NCH “BENCH”

51
Q

Cytoplasmic fixatives should ________________ which destroys mitochondria and Golgi bodies.

A

Cytoplasmic fixatives should NEVER CONTAIN GLACIAL ACETIC ACID which destroys mitochondria and Golgi bodies.

REMEMBER: Glacial acetic acid - goods na goods sa nucleus, bad na bad sa cytoplasm

**Nuclear fixatives - WITH Glacial acetic acid
**Cytoplasm fixatives - WITHOUT Glacial acetic acid

52
Q

pH of Cytoplasmic fixatives should have:

A

pH more than 4.6 (>4.6)

53
Q

Examples of Cytoplasmic fixatives:

A

Flemming’s fluid without acetic acid
Formalin with post chroming
Orth’s fluid
Helly’s
Regaud’s

mnemonic = FFOHR da Cytoplasm

54
Q

Chemical fixatives that act by creating covalent chemical bonds between protein tissue. This anchors soluble proteins to the cytoskeleton, and lends additional rigidity to the tissue/Enhance stabilization of protein

A

CROSS-LINKING FIXATIVES

54
Q

TWO MAJOR GROUPS OF CHEMICAL FIXATIVES

A
  1. CROSS-LINKING FIXATIVES (e.g., Aldehydes)
  2. PRECIPITATING (OR DENATURING FIXATIVES) (e.g., Alcoholic fixatives)
55
Q

Chemical fixatives that act by reducing the solubility of protein molecules and (often) by disrupting the hydrophobic interactions that give many proteins their tertiary structure. the precipitation and aggregation of proteins is a very different process from the crosslinking that occurs with the aldehyde fixatives/Destroys/further slow down stabilization

A

PRECIPITATING (OR DENATURING FIXATIVES)

56
Q

TWO (2) METHODS OF FIXATION

A
  1. Physical methods
  2. Chemical method
57
Q

_____ methods include HEATING, MICROWAVING, and CRYO-PRESERVATION (FREEZE-CRYING)

A

Physical methods

58
Q

_____ fixation is usually achieved by immersing the specimen in the fixative solution (IMMERSION FIXATION) or, in the case of small animals or some whole organs such as a lung, by perfusing or injecting the vascular system with fixative (PERFUSION FIXATION)

A

Chemical fixation/method

HISTOPATHOLOGY (TopnotcherQT) (26 decks)

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