Anti Streptolysin O (ASO) Test: Principle, Procedure and Interpretation

The test principle, procedure and interpretation presented here are based on Latex Anti-Streptolysin O (ASO) test.  The presence of Anti streptolysin O antibodies in serum can be indicated by seeing the ability of  serum to neutralize the hemolytic activity of streptolysin O reagent (antigen).

It is a rapid latex agglutination test for the qualitative and semi-quantitative determination of anti-streptolysin-O antibodies (ASO) in serum. In infections caused by  β-haemolytic streptococci,  streptolysin-O is one of the two hemolytic exotoxins  liberated from the bacteria that stimulates production of  ASO antibodies in the human serum.  The presence and the level of these antibodies in a serum may reflect the  nature and severity of infection.

ASO latex reagent is a stabilized buffered suspension of polystyrene latex particles that have been coated with Streptolysin O.

ASO Latex test

When the latex reagent is mixed with a serum containing ASO, agglutination occurs. The sensitivity of the latex reagent has been adjusted to yield agglutination when the level of ASO is greater than 200 IU/ml, a level determined to be indicative of disease by epidemiological and clinical studies. Sera having titers of between 200 IU/ml to 3500 IU/ml will be reactive.

Sample Collection and Handling:  Only fresh serum specimens should be used. Plasma must not be used since fibrinogen may cause non-specific agglutination of the latex. It is preferable to test samples on the same day as collected. Serum samples may be stored at 2-8°C for up to 48 hours prior to testing. If  longer storage is necessary,  sera should be stored frozen at -20°C.

Materials used in the ASO Test

  1. ASO Antigen: A stabilized buffered suspension of polystyrene latex particles coated with Streptolysin O and 0.1% sodium azide as preservative. Shake well prior to use.
  2. ASO Positive Control: Human serum containing more than 200 IU/ml ASO and 0.1% sodium azide as preservative.
  3. ASO Negative Control: Human serum containing 0.1% sodium azide as preservative.
  4. Sufficient disposable pipettes.
  5. Glass test slide.

Procedure for Antistreptolysin O Test:

  1. Bring all test reagents and samples to room temperature.
  2. Use a disposable pipette to draw up and place one free-falling drop of each undiluted sample into its identified circle of the slide. Retain each pipette for mixing in step 5.
  3. Deliver one free-falling drop of positive and negative control into its identified circle.
  4. Mix the ASO latex reagent by gently shaking. Add one free-falling drop of reagent to each control and sample.
  5. Using the flattened end of the appropriate plastic pipette as a stirrer (step 2), thoroughly mix each sample with reagent within the full area of the circle.
  6. Discard the disposable pipette.
  7. Slowly rock the slide for exactly two (2) minutes and observe for agglutination under a high intensity light.
  8. Record results.
  9. Re-wash glass slide for future use

Test Result:  A test sample is considered to contain ASO antibodies in excess of 200 IU/ml when agglutination (clumping) is observed when compared to the result of the negative control.