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Stabilization media increases recovery in paucicellular cerebrospinal fluid specimens submitted for flow cytometry
B Greig1, M Stetler-Stevenson, J Lea
1Department of Pathology, Vanderbilt University Medical Center, Nashville, Tennessee.
Insights
Stabilizing cerebrospinal fluid (CSF) with RPMI 1640 alone, without fetal calf serum (FCS), significantly increases cell yield for flow cytometric immunophenotyping (FCI). This cost-effective method improves diagnostic accuracy for hematologic malignancies.
Area of Science:
- Hematology
- Clinical Pathology
- Flow Cytometry
Background:
- Flow cytometric immunophenotyping (FCI) offers enhanced sensitivity for detecting lymphoma and leukemia in cerebrospinal fluid (CSF) compared to traditional cytomorphology.
- Low cellularity in CSF samples historically presents a challenge for accurate FCI analysis.
- Existing stabilization methods using media like RPMI with fetal calf serum (FCS) improve cell yield but increase costs.
Purpose of the Study:
- To evaluate the efficacy of RPMI 1640 without additives as a cost-effective stabilization medium for CSF samples in FCI.
- To determine if fetal calf serum (FCS) is essential for achieving high-quality FCI results in paucicellular CSF specimens.
Main Methods:
- A comparative analysis of FCI results was performed on CSF specimens stabilized with RPMI 1640 alone versus non-stabilized CSF.
- Sample processing adhered to established Clinical and Laboratory Standards Institute (CLSI) guidelines.
Main Results:
- 93% (98/105) of CSF specimens stabilized with RPMI 1640 yielded adequate viable cells (>100) for FCI.
- Only 30% (65/217) of non-stabilized CSF specimens had sufficient viable cells for analysis, with 70% being quantity not sufficient (QNS) or below viability limits.
Conclusions:
- Utilizing RPMI 1640 without FCS as a stabilization medium significantly increases cell yield for FCI of CSF.
- This approach enhances the quality of FCI results in paucicellular CSF specimens without the added expense of FCS.
Background:
Flow cytometric immunophenotpying (FCI) of cerebrospinal fluid (CSF) and other paucicellular fluids has been demonstrated to have increased sensitivity in detection of lymphoma and leukemia when compared to cytomorphology [(1) de Graaf et al., Cytometry Part B 2011, 80B:271-281; (2) Szamosi et al., CLSI Document H56-A-Body Fluid Analysis for Cellular Composition; Approved Guideline, Wayne, PA: Clinical and Laboratory Standards Institute, 2006; (3) Kraan et al., Flow Cytometric Immunophenotyping of Cerebrospinal Fluid. Current Protocols in Cytometry, Hoboken, NJ: Wiley, 2008]. However, low cellularity has been an historical problem with these samples. Several studies indicate that immediate addition of a stabilization media (e.g., RPMI with fetal calf serum (FCS)) to CSF improves the cell yield for FCI [(1) de Graaf et al.]. Such stabilization medias can, however, significantly increase cost.
Methods:
We compared FCI results in CSF stabilized with RPMI 1640 (without additional additives) to results obtained using non-stabilized CSF. Samples were processed according to published CLSI guidelines [(2) Szamosi et al.].
Results:
About 98/105 (93%) CSF specimens stabilized with RPMI had adequate numbers of viable cells (>100) for performing FCI. About 65/217 (30%) CSF specimens without stabilization had adequate numbers of viable cells for analysis (70% either quantity not sufficient (QNS) or specimen viability below analytical limits).
Conclusions:
Utilizing RMPI without FCS as a stabilization media results in increased cell yield and improved FCI results. We have found FCS is not required to achieve high quality results in FCI of paucicellular CSF specimens.
