C3d-containing Circulating Immune Complex
Enzyme-linked immunosorbent assay method for the semi-quantitative determination of specific C3d-containing circulating immune complexes in human serum.
Circulating immune complexes are present in many individuals with Systemic Lupus Erythematosus (SLE) and Rheumatoid Arthritis (RA), especially with any of the vasculitides complications. Levels of CICs have been reported to show correlation with disease activity in that higher levels are reported during active phases of the disease.
Many tests have been developed for the detection of CICs, including PEG precipitation and radial immunodiffusion. No single procedure appears to detect all types of CIC however, those procedures, such as the Raji cell assay and C3d solid phase assay, which detect CICs containing fragments of C3 (i.e. C3d) appear to detect clinically relevant events.
The AutostatII test system for C3d circulating immune complexes detects immune complexes containing both C3d and IgG. The concentration is expressed as μg/ml heat aggregated human globulin (HAG) equivalents.
The AutostatII test range also includes a kit for C1q circulating immune complexes which detects immune complexes containing both C1q and IgG.
The AutostatII assay is a solid phase immunosorbent assay (ELISA) in which the analyte is indicated by a colour reaction of an enzyme and substrate. The AutostatII wells are coated with anti-C3d monoclonal antibody (1).
On adding diluted serum to the wells the C3d-containing CICs (2) present bind to the antibody. After incubating at room temperature and washing away unbound material, horseradish peroxidase conjugated anti-IgG monoclonal antibody (3) is added, which binds to the immobilised complexes.
Following further incubation and washing, tetra-methyl benzidine substrate (TMB) (4) is added to each well. The presence of the complex turns the substrate to a dark blue colour. Addition of the stop solution turns the colour to yellow.
The colour intensity is proportional to the amount of immune complexes present in the original serum sample.