The document describes the Ascoli test, a precipitin test used for the serological diagnosis of anthrax. The test uses antiserum raised in rabbits against Bacillus anthracis to detect thermostable antigens from the bacteria. A positive result is indicated by the formation of a visible precipitate band within 15 minutes when the antiserum is layered with a filtrate obtained from boiling and filtering a sample suspected to contain B. anthracis antigens.

2. Ascoli Test
Abdul Wahab 15-arid-4445

3. Introduction:
• In 1911 Ascoli published a procedure for the detection of thermostable
antigen of anthrax in animal tissues.
• A precipitin test (a test in which an antibody produces a visible precipitate
when it reacts with its antigen) used in the serological diagnosis of
anthrax.

4. Anthrax
• Bioterrorism importance
• Worldwide in distribution
• Caused by Bacillus anthracis
• Gram positive cylindrical rods
• Aerobic and spore forming organisms

5. Ascoli Test:
• The test uses antiserum raised in rabbits to produce a precipitin reaction
• It lacks high specificity in that the thermostable antigens of B.anthracis
are shared by other bacillus species and it is dependent on that , only
B.anthracis would proliferate throughout the animal and deposit
sufficient antigen to give positive reaction.

6. Conti…
Antiserum production:
• Antiserum is prepared in rabbit by subcutaneous inoculation of Sterne
Anthrax vaccine on day 1 and 14
• On day 28 and 35 rabbits are administered 0.5 ml of a mixture of several
strains of virulent B.anthrax
• Live virulent bacteria can be inactivated by prolonged suspension in 0.2%
formalized saline

7. Conti…
• Formalized suspension dose is increasing from 0.1, 0.5, 1 and 2 ml I/V at
intervals of 4-5 days after initial vaccination on day 1 and 14
• Following either procedure a bleed at 10 days after the last injection
should determine whether additional 2 ml doses should be administered to
boost the precipitin

8. Procedure:
• Put approximately 2 g of sample(usually blood) in 5 ml of saline containing
1:100 final concentration of acetic acid
• Boil for 5 minutes
• The resultant solution is cooled
• Cooled solution is filtered through filter paper and a filtrate is obtained
• A few drops of rabbit antiserum are placed in small test tube
• Filtered solution (filtrate) is gently layered over the top of antiserum

9. Results:
• A positive test is the formation of a visible precipitin band in under 15
minutes otherwise test is negative.