Aim: To demonstrate calcium in paraffin sections.

Principle: Tissue sections are treated with silver nitrate solution, the calcium is reduced by the strong light and replaced with silver deposits, visualized as metallic silver.

Reagents:-

5% silver nitrate solution
Silver nitrate                       25gm
Distilled water                    500ml

Mix well, pour into acid bottle. Store in refrigerator (stable for one year).

Control: Positive control

Procedure:-

Bring sections to water. One section works as negative control.
Immerse one section in citrate buffer (pH4.5) for 20 minutes to remove
calcium if it is present.
Wash both the slides in distilled water.
Put 5% silver nitrate solution.Expose the slides to bright sun light for 10 to 20 minutes.
Wash in distilled water several times.
Treat with 5% sodium thiosulfate for 2 to 3 minutes.
Wash well in water.
Counter-stain with neutral red or Van Gieson stain.
Dehydrate, clear in xylene and mount in DPX.

Result:-

Calcium                             Dark green or black
Back ground                     Depends on counter stain.

Note: Silver nitrate is tumorogenic and oxidizer. It is strong skin and eye irritant.