Plasma IgM mediates complement activation by PF4/heparin complexes. (A) Commercial IgM (0-1000 μg/mL; filled symbols) or IgG (0-5000 μg/mL; open symbols) or monoclonal myeloma IgM (0-1000 μg/mL; hatched symbols) was added to the plasmas of 2 donors with a low complement response type (circle/square), and complement activation by PF4/heparin was measured by the antigen-C3c capture ELISA assay. Graph shows complement activation (y-axis) as a function of added immunoglobulin concentration. (B) Plasma with an intermediate donor phenotype was incubated with anti-IgM or control beads, followed by the addition of buffer, PF4/heparin, or PF4/heparin +400 µg/mL IgM, and complement activation was measured by the antigen-C3c capture ELISA assay. Graph shows complement activation (y-axis) in control or IgM depleted plasma (x-axis). (C) Plasma with a low donor phenotype was incubated with varying antigen concentrations (PF4; 0-25 μg/mL + heparin; 0-0.25 U/mL) and IgM (0-800 μg/mL), and complement activation was measured by the antigen-C3c capture ELISA assay. Graph shows complement activation response at varying IgM concentrations (y-axis) as a function of PF4/heparin concentrations (x-axis). *P < .005; **P < .0001. Results are shown from a representative experiment involving 3 donors tested on 3 different occasions.