LBOR02: CROSSMATCH OPTIONS? CAN A CELL CAPTURE IMAGE BE WORTH A THOUSAND FLOW EVENTS?

Abstract

Aim There have been no major changes in reagent or instrumentation in the flow cytometry crossmatch (FC-XM) since its description in 1983. The cost of the FC instrumentations, calibrations and maintenance is a restraint for many histocompatibility labs. In this study we evaluated the Cellometer Image Cytometer as an alternative instrument for the FC-XM. Methods Our FC-XM protocol was modified to a 2-color panel for T and B cells. Each panel consists of Neg. and Pos. control, and the sample. After the initial incubation, a cocktail consisting of anti CD3-PeCy5/anti-F(ab’)2-PE and anti CD19-PeCy5/anti-F(ab’)2-PE was added to the T and B cell panel respectively. The sample was added to a counting chamber that allowed the formation of a cellular monolayer. Images were captured using the Cellometer Vision CBA, an image-based cytometer designed for cell counting, size, and fluorescence intensity detection. On Fig. 1 we show one of the multiple pictures obtained for the cell plot analysis and the shift of the PE channel. A 2-SD cut off was generated for the T and B cell XM analysis. Results On this study we have tested 31 T and B cell XM on the Cellometer to compare with our standard 3C-FCXM. We obtained a concordance of 94% and 100% for T and B cells XM respectively. The Fisher exact test p value was Conclusion This feasibility study demonstrates that the FC-XM reaction could be easily adapted to the Cellometer without compromising specificity and sensitivity. The low instrumentation cost, minimal maintenance, calibration, and operator training are among the advantages of this instrument. With these advantages, and the results obtained here, we demonstrate that the Cellometer Vision CBA is an attractive option for the histocompatibility laboratories.