Determination of plasma C3a des Arg levels after blood contact with foreign surfaces

The products of complement activation, including C3a, C4a, C5a, and their des Arg derivatives, have been used by many investigators as indicators of blood or foreign surface bioincompatibility, e.g., during hemodialysis. One problem encountered is that after extracorporeal circulation over some dialysis membranes very high C3a levels (5,000-10,000 ng/ml) are generated. To the authors' knowledge the only presently available C3a des Arg RIA kit is provided by Upjohn Diagnostics (Kalamazoo, MI, U.S.A.), which covers an assay standard range of 20-500 ng/ml. Since Upjohn's assay procedure manual does not contain a recommended dilution technique the authors undertook a study to determine the best method. The diluents tested include physiologic saline, normal plasma, phosphate-buffered saline-gelatin, and the Upjohn precipitating agent provided in their kit. The results show that only dilution by normal plasma gave reliable results. The use of the other three diluents resulted in an overestimation of C3a levels. Therefore it is important to know the dilution technique used when comparing the results of membrane C3a generating ability presented by different authors. Furthermore, comparison of different membranes where plasma samples in some cases do not require dilution (e.g., polyacrylonitrile membranes) with those requiring dilution (e.g., regenerated cellulose membranes) will lead to distorted results if an accurate technique is not used.