Decalcification

Question 1

Removal of calcium and lime salts

Answer 1

Decalcification

Question 2

Done after fixation and before dehydraation and impregnation

Answer 2

Decalcification

Question 3

Calcium might interfere with accurate evaluation and examination

Answer 3

Decalcification

Question 4

Facilitate normal cutting of tissue in sectioning

prevent obscuring microanatomical detail of
tissue

Answer 4

Decalcification

Question 5

Organs that requires decalcification

Answer 5

1. Bones - because calcium is present in bones
2. Tuberculous lungs - calcified lungs because of
   TB bacteria
3. Atherosclerotic vessels - lipids, calcium,
   cellular debris in the lumen of blood vessels
4. Teeth

Question 6

FACTORS AFFECTING THE RATE OF
DECALCIFICATION

Answer 6

1. Concentration
   ○ If less concentrated, slow-acting
2. Tissue-to-volume ratio
   ○ 1:20 (tissue:decalcifying agent or
   solution)
3. Temperature
   ○ Must be regulated
4. Mechanical agitation
   ○ Key to a rapid decalcifying of sample
5. Size and consistency of tissue sample
   ○ If sample is thick and rigid, longer
   decalcifying time

Question 7

Four types of decalcifying agents

Answer 7

1. Acid
2. Chelating agents
3. Ion Exchange resin
4. Electrophoresis

Question 8

Routine or most common decalcifying agent

Answer 8

Nitric acid (5-10%)

Question 9

Imparts yellow coloration d/t nitrous acid formation

Answer 9

Nitric acid (5-10%)

Question 10

also acts as tissue
softener; Nitric acid + Chromic acid +
ROH

Answer 10

Perenyi’s fluid

Question 11

Fastest agent (simple or compound) 5-10%

Answer 11

Phloroglucin Nitric acid

Question 12

Provides good nuclear staining at 1%

Answer 12

Hydrochloric acid

Question 13

Slower and causes more distortion compared to HNO3

Answer 13

Hydrochloric acid

Question 14

For teeth and small bones

Answer 14

Von Ebner’s fluid

Question 15

HCI + 36% NaCH + distilled water

Answer 15

Von Ebner’s fluid

Question 16

For routine decalcification of post-morten research tissues, small pieces of bone and teeth

Answer 16

Formic acid

Question 17

Tissue sample is from human who is already deceased

Answer 17

Post-mortem

Question 18

Good for in situ hybridization staining

Answer 18

Formic acid

Question 19

is better than HNO3

Answer 19

Formic acid - sodium citrate solution

Question 20

Small bone spicules

Answer 20

Trichloroacetic acid

Question 21

Good nuclear staining

Answer 21

Trichloroacetic acid

Question 22

Minute pieces of bone

Answer 22

Sulfuruous acid

Question 23

Minute bone spicules

Answer 23

Chromic acid (flemming’s fluid)

Question 24

Use of salts to form complexes with catt salets for ease of removal

Answer 24

Cheating agents

Question 25

Immunohistochemistry % enzyme staining

Electron miscroscope

Answer 25

Cheating agents

Question 26

Duration: 1-3 weeks for small spx

6-8 weeks for longer & dense bones

Answer 26

Cheating agents

Question 27

pH: 7-7.4

Answer 27

Cheating agents

Question 28

Common brand of cheating agents

Answer 28

Cal ex
Versene

Question 29

Mainly removing calcium ions from from formic
acid containing decalcifying solutions

Answer 29

Ion exchange resin

Question 30

Increase solubility (uses formic acid with TSE:Formic acid of 1:20-30)

Answer 30

Ion exchange resin

Question 31

● Ca++ moves to cathode
● P: small bone fragments

Answer 31

Electrophoresis

Question 32

Electrophoresis uses __% acid

Answer 32

88% formic acid