A flow cytometric method for detection and enumeration of low-level, residual red blood cells in platelets and mononuclear cell products

Abstract

BACKGROUND: Conventional automated cell counters cannot accurately count residual red blood cells (rRBCs) that are often present in various blood products. A two-color flow cytometric method (FC) was validated for detecting and enumerating rRBCs in platelets (PLTs) and mononuclear cell (MNC) products. STUDY DESIGN AND METHODS: PLT and MNC products for PLTs (CD61-fluorescein isothiocyanate) and rRBC (anti-glycophorin A-phycoerythrin) were double stained, and data were acquired with a flow cytometer. Assay linearity, accuracy, and precision were assessed with a standard-dilution series of rRBCs. This assay was used to determine the rRBCs of apheresis PLTs collected with Trima Accel (Gambro BCT) and MNC products collected with the COBE Spectra (Gambro BCT). RESULTS: The linear range of this assay in PLT and MNC products is 10 to 2000 RBCs per μL (R2 = 0.994). FC had a mean intraassay coefficient of variation of 11.8 percent at 34 RBCs per μL. A standard clinical hematology analyzer overestimated rRBCs in MNC products by 1.59 × 105 ± 0.7 × 105 RBCs per μL. Apheresis PLTs had a median of 17.4 RBCs per μL, with 99.0 percent containing fewer than 90.0 RBCs per μL. CONCLUSIONS: This method for determining rRBCs in blood products is accurate and repeatable with a lower limit of detection adequate to assess currently available blood products. FC should be considered for determining rRBCs in MNC products.