DECALCIFICATION

SYNOPSIS:
Decalcification
Decalcifying agents
Acids
Chelating agents
Factors affecting decalcification
End point of decalcification

1)DECALCIFICATION:
• It is the process of removal of calcium from the organic collagen matrix, calcified cartilage
and surrounding tissues.
• Carried out in chemical agents called as DECALCIFYING AGENTS.

2) Decalcifying agents
Good decalcifying agent is the one
• Which removes all the calcium from tissue completely
• Does not cause damage to the cells and fibres
• Does not interfere in staining technique
• Gives fast decalcified material

Two types
1)Acids and
2) Chelating agents
2a) ACID DECALCIFIERS:
Divided into :
a) Organic [weak] acids.
Eg: Formic acid

b) Inorganic [strong]acids.
Eg:
Hydrochloric acid:
Sulfurous acid:
Nitric acid:

2b) CHELATING AGENTS:

EDTA

These binds to ionized calcium on the outside of apatite crystal.

Calcium from inner surface replaces the outer.
Crystal size becomes smaller.
Advantages: Do not damage the tissue
Good stainability of the tissue
Specific PH can be adjusted(immunochemistry)
Disadvantages: Prolonged duration.
Most commonly used agent is the EDTA/ethyelenediaminetetraceticacid.

3) FACTORS AFFECTING DECALCIFICATION:

Concentration of decalcifying agent
Temperature
Agitation
Suspension
Other factors such as age of the patient, type of bone, size of the specimen , concentration and
volume of the active reagent also affect the process.

COMPLETION OF DECALCIFICATION:
Done by decalcification end point test–to know when calcium removal is complete.
Formic acid, Hydrochloric acid and nitric acid use requires daily testing and near end point it is
necessary to test every five hours.
EDTA use requires weekly test.

4) DECALCIFICATION END POINT:

METHODS INCLUDES:
Physical - Probing, needling, slicing, bending or squeezing tissue
Chemical - Calcium oxalate test
Radiographic method