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HCT > Lec 6: Decalcification > Flashcards

Lec 6: Decalcification Flashcards

1
Q

Enumerate the steps for Conventional Tissue Processing

A
  • Fixation
  • Decalcification
  • Dehydration
  • Clearing
  • Infiltration
  • Embedding
  • Trimming/Cleaning
  • Sectioning
  • Staining
  • Mounting
  • Labeling
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2
Q

The removal of calcium ions from a bone or calcified tissue through a histological process.

A

Decalcification

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3
Q

The process which makes the bone or calcified tissue flexible and easy to cut.

A

Decalcification

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4
Q

Tissues that include Decalcification

A
  • bones
  • teeth
  • calcified tumor
  • calcified heart valve
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5
Q

The substance that makes the bone hard

A

Calcium

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6
Q
  1. It forms soluble calcium salts in anion exchange which moves calcium from the bone into the decalcifying solution.
  2. It sequesters metallic ions including calcium in aqueous solutions.
A
  1. Strong Mineral Acid or Weak Organic Acid
  2. Chelating agent
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7
Q

Criteria of a good Decalcifying Agent

A
  • Complete removal of calcium
  • absence of damage to tissue cells or fibers
  • Non impairment of subsequent staining technique
  • Reasonable speed of the calcification
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8
Q

What are the factors affecting the rate of decalcification (5)

A
  • concentration
  • fluid access
  • size & consistency
  • agitation
  • temperature
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9
Q

Factors affecting Decalcification Rate:

[TRUE or FALSE]
1. Increased temperature = slows down Decalcification = increases tissue damage
2. The bigger the tissue, the longer the decalcifying period.
3. The denser the tissue, the faster the decalcifying period.
4. Higher concentration = slower Decalcification

A
  1. FALSE
  2. TRUE
  3. FALSE
  4. FALSE
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10
Q

Factors affecting Decalcification Rate:

  1. Higher concentration does what to the tissue?
  2. The optimum temperature recommended.
  3. Recommended Fluid to Tissue ratio
  4. How does gentle agitation & low-speed stirring/rocking affect Decalcification Rate?
  5. How long do dense bone tissues usually take to complete Decalcification?
A
  1. Decalcifies rapidly, but more harm to tissue
  2. RT (18–30°C)
  3. 20:1
  4. Increase Decalcification Rate
  5. 14 days +
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11
Q

Procedure of Decalcification

A
  1. Selection of tissue
  2. Fixation
  3. Decalcification
  4. Acid Neutralization
  5. Thorough washing
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12
Q

In tissue selection, what is the ideal thickness for:
a.) dense bone
b.) soft bone

A

a.) 2 to 5 mm
b.) 4 to 6 mm

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13
Q

When cutting large specimen:

1.) What saw is used?
2.) What saw is NOT advisable to use?

A
  1. Large fine-toothed bone saw
  2. Coarse Saw
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14
Q

When cutting a small specimen, what is used?

A

Geological cutting machine with diamond impregnated cutting disc

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15
Q

It protects the cellular elements of the bone from damage caused by acids in decalcifying agents.

A

Fixation

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16
Q

FIXATION:

  1. Fixative for EM
  2. Fixatives for good preservation of Bone Marrow
  3. Fixatives for Tooth specimens
  4. Fixatives for Bone
A
  1. Glutaraldehyde
  2. a.) Zinc formalin mixture
    b.) B-5, formol-acetic alcohol (Davidson’s solution)
    c.) Bouin’s solution
  3. 15% formic acid
  4. Buffered formalin
17
Q

The three main types of decalcifying agents

A
  • strong mineral acids
  • weak organic acids
  • chelating agent
18
Q

The 2 methods of Decalcification

A
  1. Acid method
    » strong mineral acids
    » weak organic acids
  2. Chelation
    » chelating agent
19
Q

Strong Mineral Acid:

  • most common
  • fastest
  • recommended for routine purposes
  • at higher concentration, inhibits nuclear stain & damages tissue
A

Nitric Acid

20
Q

NITRIC ACID:

  • Decalcification time: 12 to 24 hours
  • easily remove by 70% alcohol
  • recommended for urgent biopsy
  • produces good nuclear staining
  • can be used for large or heavily mineralized cortical bone specimen
A

Aqueous Nitric Acid Solution 10%

21
Q

NITRIC ACID:

  • Decalcification time: 12 to 24 hours
  • imparts a yellow color (impairs staining reaction)
  • damages tissue & tissue antigens
A

Aqueous Nitric Acid Solution 10%

22
Q

NITRIC ACID:

  • Decalcification time: 1 to 3 days
  • less tissue damage than 10% Aqueous Nitric Acid
  • recommended for urgent biopsy
  • good nuclear staining
  • imparts a yellow color
A

Formol-Nitric Acid

23
Q

NITRIC ACID:

  • Decalcification time: 2 to 7 days
  • decalcifies and softens tissue
  • maceration is avoided (presence of chromic acid & alcohol)
  • recommended for routine purposes
  • good Nuclear & Cytoplasmic staining
  • slow for dense bones
A

Perenyi’s fluid

24
Q

NITRIC ACID:

  • Decalcification time: 12 to 24 hours
  • most rapid
  • poor Nuclear staining
  • imparts yellow color
A

Phoroglucin-Nitric Acid

25
Q
  • does NOT require washing out before dehydration
  • recommended for teeth and small pieces of bone
A

Von Ebner’s Fluid

26
Q

FORMIC ACID:

  • Decalcification time: 2 to 7 days
  • fixative & decalcifier
  • excellent Nuclear & Cytoplasmic staining
  • recommended for teeth & small pieces of bone
  • for most routine surgical specimens
  • slow
  • requires Neutralization (5% Sodium suilfate & wash)
A

10% Formic Acid

27
Q

FORMIC ACID:

  • Decalcification time: 3 to 14 days
  • slow
  • better nuclear staining than Nitric Acid
  • recommended for autopsy materials, bone marrow, cartilage and tissue studies for research purposes
A

Formic acid - sodium citrate solution

28
Q
  • Decalcification time: 4 to 8 days
  • good nuclear staining
  • does NOT require washing out
  • NOT used for dense tissues
  • only for small spicules of bone
A

Trichloroacetic acid

29
Q
  • a fixative & decalcifying agent
  • for minute bone spicules
  • inhibits nuclear staining
  • forms precipitate
  • forms insoluble pigments (dehydration)
  • environmental toxin & carcinogenic
A

Chromic acid (Flemming’s Fluid)

30
Q

Chromic acid

  1. pH
  2. Decalcification time
A
  1. 4.5
  2. 6 days
31
Q

Most common chelating agent in the market

A

EDTA

32
Q

EDTA:

  1. Action of EDTA
  2. Commercial name
  3. Duration of tissue in EDTA
    a. Small specimens
    b. Dense cortical bone
  4. Solution should be changed every ___ days.
A
  1. Captures and combines with calcium to form an insoluble non-ionized complex.
  2. Versene
  3. a. 1 to 3 weeks
    b. 6 to 8 weeks or longer
  4. Every 3 days
33
Q

The effects of the following to the tissue:
1. Over Decalcification
2. Under Decalcification

A
  1. Over Decalcification
    • spoils the staining of basophilic elements
    • causes maceration of the softer tissue elements
  2. Under Decalcification
    • interferes with proper sectioning & staining
34
Q

It is the process where the chemicals used in decalcification will be neutralized in order to prevent a reaction of the chemicals to the succeeding pathologic processes

A

Acid Neutralization

35
Q

The chemical solutions used to accomplish Chemical neutralization

A
  • saturated lithium carbonate solution
  • 5 to 10% aqueous sodium bicarbonate solution
36
Q

How long are the following rinsed in tap water?

a.) small sample
b.) larger specimens

A

a.) 30 minutes
b.) 1 to 4 hours

HCT (9 decks)

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