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SOP

Anti-Streptolysin O (ASO) Testing

SOP for ASO titer by latex agglutination for evidence of recent group A streptococcal infection and post-streptococcal sequelae.

Last verified 1 month ago 3 min read Immunology Serology / Immunology #aso #antistreptolysin #strep #rheumatic fever #serology
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Document Control
Version
1.0
Effective
Aug 30, 2026
Reviewed
Aug 30, 2026
Department
Serology / Immunology
Principle

Streptolysin O is an oxygen-labile hemolysin produced by group A streptococci. Infection triggers an IgM/IgG antibody response measured as the ASO titer. Agglutination of streptolysin O-coated latex particles increases with the amount of antibody, allowing semi-quantitative titering; the peak titer lags weeks behind infection, so serial sampling captures the rise.

Standard Operating Procedure: Anti-Streptolysin O (ASO) Testing

1. Purpose and Scope

To detect and quantitate antibody to streptolysin O (ASO) as evidence of a recent infection by group A (and some other group) beta-haemolytic streptococci, useful in the evaluation of rheumatic fever, post-streptococcal glomerulonephritis, and other suspected post-streptococcal sequelae.

2. Specimen Requirements

  • Type: Serum (plain tube or SST).
  • Volume: Minimum 0.5 mL.
  • Interference: Lipemic, icteric, or grossly hemolyzed serum may interfere with turbidimetric reading.
  • Timing: ASO titers rise 1-3 weeks after infection and peak at approximately 3-6 weeks; a single acute sample may be falsely low, so serial (paired) samples are preferred when post-streptococcal disease is suspected.

3. Safety and Precautions

  • Standard PPE (gloves, lab coat). Serum is potentially infectious.
  • Dispose of sharps and biohazard waste per laboratory policy.
  • Do not mouth-pipette.

4. Equipment and Reagents

  • Latex agglutination slide or automated turbidimetric analyzer.
  • ASO latex reagent (streptolysin O-coated latex particles) and positive/negative controls.
  • Isotonic saline or reaction buffer.

5. Step-by-Step Procedure

  1. Allow reagents and serum to come to room temperature; mix latex reagent gently (do not shake rigorously to avoid foam).
  2. Run positive and negative controls before patient samples each day.
  3. For semi-quantitative titer: prepare twofold dilutions of serum (e.g., 1:20 to 1:1280), place one drop of each dilution on the slide, add one drop of latex reagent, and rotate; read agglutination at the endpoint.
  4. For automated turbidimetric/nephelometric assay: load serum, run the calibration curve and controls, and report the quantitative concentration and the correlated titer.
  5. Record the reportable titer or concentration; for paired sera, report the acute and convalescent values together.

6. Quality Control

  • Positive and negative controls with each run; lot-to-lot validation of the latex reagent.
  • Enforce Levey-Jennings rules for the automated method; participate in EQA.
  • Document the endpoint reading criteria for semi-quantitative methods.

7. Decision limits

The interpretive thresholds below are clinical decision limits; titers are read as the highest dilution showing agglutination.

  • Significant elevation: titers above the laboratory's pediatric/adult upper limits (commonly 200-320 IU/mL or a 1:200 titer) indicate recent streptococcal infection. Children typically have higher normal titers than adults.
  • Paired sera: a fourfold (two-tube) rise between acute and convalescent (2-4 weeks apart) samples is the strongest evidence of recent streptococcal infection; a single elevated titer is supportive but less definitive.
  • Timing: false negatives occur when only a very early acute sample is tested, before antibodies have risen.
  • Limitations: ASO does not prove causation of rheumatic fever or glomerulonephritis; it indicates a recent immune response. Anti-DNase B may be added for skin (impetigo) streptococcal infections where ASO response can be weak.

Frequently Asked Questions

ASO rises weeks after infection; a single acute sample can be below the threshold. A fourfold rise between acute and convalescent samples firmly documents recent streptococcal infection.
Skin (impetigo) streptococcal infections often produce a weak ASO response. Adding anti-DNase B antibody testing increases sensitivity for cutaneous infections.
No. A high titer only confirms a recent immune response to streptococcus; the diagnosis of acute rheumatic fever uses clinical criteria (Jones) supported by the serologic evidence.