New procedure for the detection of complement deficiency by ELISA. Analysis of activation pathways and circumvention of rheumatoid factor influence.
A procedure using enzyme-linked immunosorbent assays for the assessment of complement function has been evaluated. The sera investigated were incubated in microtiter plates with solid-phase complement activators. Human polyclonal IgG or monoclonal IgM were used for classical activation pathway assays and Salmonella typhosa lipopolysaccharide (LPS) for alternative activation pathway assays. The analysis focussed on deposition of C9 and properdin as detected with enzyme-conjugated antibodies. In an attempt to avoid spurious results due to rheumatoid factors in patient sera, monoclonal mouse and chicken antibodies were unsuccessfully tested as indicator reagents in the assay with solid-phase IgG. However, the use of solid-phase IgM as an activator completely circumvented the influence of rheumatoid factors. With solid-phase IgG or IgM, properdin deposition occurred in the absence of factor D. A combination of assays is suggested for diagnostic purposes: IgM-coated plates with detection of bound C9 and properdin for the classical pathway and LPS-coated plates with detection of bound properdin for the alternative pathway. The procedure distinguished between defects of the classical activation pathway (C1, C4, C2), the alternative activation pathway (C3, factor B, factor D, properdin) and the terminal components (C5-C9). This analytical approach may be useful for detection of inherited complement deficiency and the assessment of complement function in acquired complement deficiency states.
Fredrikson
G N
GN
Department of Medical Microbiology, Clinical Immunology Section, University of Lund, Sweden.
Truedsson
L
L
Sjöholm
A G
AG
D003168:Complement Fixation Tests; D003170:Complement Pathway, Alternative / Q000187:drug effects; D003171:Complement Pathway, Classical / Q000187:drug effects; D003165:Complement System Proteins / Q000032:analysis / Q000172:deficiency*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007074:Immunoglobulin G / Q000494:pharmacology; D007075:Immunoglobulin M / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D015203:Reproducibility of Results; D012217:Rheumatoid Factor / Q000032:analysis*
D007074:Immunoglobulin G; D007075:Immunoglobulin M; D008070:Lipopolysaccharides; D003165:Complement System Proteins; D012217:Rheumatoid Factor