RT-PCR, FACS, and immunoprecipitation analysis of EPCR in neutrophils. (A) EPCR mRNA in neutrophils and monocytes. 1 μg total RNA from each sample was reverse transcribed into cDNA and amplified for the EPCR gene using PCR. EPCR is represented by the 409–base pair product. (B) FACS analysis of anti–EPCR mAb binding to neutrophils. Fluorescence analysis used a FACScan Flow cytometer, and a histogram of PE fluorescence is shown. Cells were either incubated with isotype-matched control IgG (black curve) or anti–EPCR mAb (gray line) and stained with PE-conjugated streptavidin. (C) Synthesis of EPCR in neutrophils. Cells were radiolabeled for 40 minutes, followed by lysis and immunoprecipitation with anti-EPCR antibody RCR-2, SDS-PAGE, and autoradiography.