How must synovial fluids be handled prior to cell count and cytospin preparation to prevent clotting and preserve sample integrity?

Handling of Synovial Fluid Prior to Cell Count and Cytospin Preparation

Synovial fluid must be collected in EDTA tubes (rather than heparin or plain tubes) to prevent clotting and preserve cellular integrity for accurate cell counts and cytospin preparations.

Rationale for EDTA Use in Synovial Fluid Analysis

EDTA (ethylenediaminetetraacetic acid) is the preferred anticoagulant for synovial fluid analysis for several important reasons:

• EDTA prevents clotting of synovial fluid which would otherwise lead to inaccurate cell counts 1
• EDTA better preserves cellular morphology compared to heparin or no preservative 1
• EDTA-preserved samples maintain stable leukocyte counts for up to 24 hours (ICC = 0.985), allowing for delayed processing if needed 1

Critical Timing Considerations

Timing is crucial when handling synovial fluid samples:

• Prompt examination is essential as leukocyte counts decrease within hours of collection 2
• Borderline inflammatory fluids (3,150-6,200 WBC/mm³) can decrease to non-inflammatory ranges (<2,000 WBC/mm³) within 5-6 hours, potentially leading to misdiagnosis 2
• Crystal identification is time-sensitive:
  • Calcium pyrophosphate dihydrate (CPPD) crystals may dissolve completely within 3-8 weeks 2
  • Monosodium urate crystals become smaller, less birefringent, and less numerous over time 2

Processing Techniques for Optimal Results

For viscous synovial fluid samples:

• Add hyaluronidase (150 USP units/ml) to break down hyaluronic acid and reduce viscosity 3
• This is particularly important for non-inflammatory synovial fluids where cell counts are low 3
• Hyaluronidase treatment followed by cytospin preparation improves cellular morphology and differential count accuracy 3

Procedural Steps for Proper Handling

1. Collect synovial fluid in EDTA tubes immediately after aspiration
2. Process the sample within 1 hour of collection whenever possible
3. For viscous samples, add hyaluronidase before cytospin preparation
4. Perform cell count and differential promptly to avoid artifactual changes
5. If processing must be delayed, refrigeration at 4°C can slow cellular degradation, but cannot prevent it completely 2

Common Pitfalls to Avoid

• Delayed processing leading to decreased WBC counts and potential misclassification of inflammatory status 2
• Using inappropriate collection tubes (plain tubes will allow clotting)
• Failure to address high viscosity in non-inflammatory fluids, which can impair accurate cell counting 3
• Misinterpretation of artifactual crystals that may form over time in stored samples 2

By following these protocols, the accuracy of cell counts and cytospin preparations will be maximized, leading to more reliable diagnosis and appropriate treatment decisions for this 2-year-old child with suspected septic arthritis.