CSF Collection Tube Order During Lumbar Puncture
The standard order for collecting CSF samples during lumbar puncture is: Tube 1 for chemistry and serology, Tube 2 for microbiology (Gram stain and culture), Tube 3 for cell count with differential, and Tube 4 (if needed) for additional studies such as cytology or special tests.
Rationale for Collection Order
The traditional teaching prioritizes placing the cell count tube last (Tube 3) because:
- The first tube collected may contain blood from the needle puncture itself, which could falsely elevate red and white blood cell counts 1
- By the third tube, any traumatic blood contamination from the procedure typically clears, providing the most accurate cell count 1
- The middle tube (Tube 2) is optimal for microbiology because it balances minimal traumatic contamination with adequate volume for culture 2, 3
Specific Tube Allocation
Tube 1: Chemistry and Serology
- CSF glucose with concurrent plasma glucose measurement 2, 3
- CSF protein concentration 2, 3
- Storage for potential serological testing if initial workup is non-diagnostic 2
Tube 2: Microbiology
- Gram stain (sensitivity 50-99%, specificity 97-100%) 3
- Bacterial culture (diagnostic in 70-85% of untreated cases) 3
- The first 2-2.5 mL should be allocated for microscopy, culture, and sensitivities 1
Tube 3: Cell Count and Differential
- Total white blood cell count with differential 1, 3
- Red blood cell count 1
- A minimum of 1-2 mL is required, with 5 mL being optimal for comprehensive testing 1
Tube 4: Additional Studies (as indicated)
- Viral PCR studies (requires additional 2 mL) 1, 3
- Tuberculosis testing (requires additional 6 mL) 1
- Cytology or other specialized tests 1
Volume Requirements
- A total collected volume of at least 5 mL is recommended for standard diagnostic procedures to avoid false-negative findings 1
- For comprehensive testing, at least 22 mL can be safely removed from adults, as CSF is produced at approximately 15 mL/hour 2, 1, 3
- Insufficient volume collection is a common cause of false-negative results in CSF analysis 1
Critical Processing Considerations
- CSF samples should be processed within 30-60 minutes of collection for optimal results 1
- Delay in processing beyond 30 minutes can lead to cellular degradation and affect test accuracy 1
- CSF should be collected in dedicated sterile collection tubes to avoid contamination 1
Essential Documentation
When performing lumbar puncture, the following must be documented:
- CSF opening pressure (unless performed in sitting position) 2, 3
- CSF appearance (clear, turbid, cloudy, purulent, or bloody) 3
- Concurrent plasma glucose at time of CSF collection 2, 3
Common Pitfall to Avoid
Do not assume a traumatic tap invalidates the cell count. For traumatic taps, white cell counts can be approximately corrected by subtracting 1 white cell for every 700-7000 red blood cells in the CSF 1. However, collecting the cell count in the third or fourth tube minimizes this issue by allowing traumatic blood to clear through the earlier tubes.