Methodological pitfalls in serum IgG2 level measurements by immunoenzymatic assays with monoclonal antibodies.
Four anti-IgG2 monoclonal antibodies (Mabs) were evaluated for their reactivity with purified myeloma IgG2 of different light chain types and Gm allotypes in three distinct immunoenzymatic assays (ELISA). The reactivity of three Mabs with solid-phase antigens was similar whereas an anti-Fab antibody (clone HP 6114) predominantly bound IgG2 kappa. In competitive and immunometric (sandwich type) assays, the binding of the two anti-IgG2 Mabs (HP 6014 and HP 6114) reacting with epitopes located on the Fab fragment was strongly influenced by the light chain type of IgG2 and by other factors (probably including differences in the variable regions); the Mab HP 6114 reacted virtually only with IgG2 kappa whereas the Mab HP 6014 displayed a much stronger affinity for IgG2 lambda than for IgG2 kappa; for both anti-Fab Mabs, important differences were found in their binding to individual IgG2 proteins. In addition, the Mab HP 6014 seemed to show a slightly better affinity for IgG2 bearing the Gm(23) allotype. These results urge much caution in IgG2 level measurement, especially with commercial kits, most of which use the Mab 6014 as the single anti-IgG2 reagent.
Aucouturier
P
P
Laboratory of Immunology and Immunopathology, CNRS URA 1172, CHUR, Poitiers, France.
Lacombe
C
C
Preud'homme
J L
JL
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D000939:Epitopes; D005069:Evaluation Studies as Topic; D006801:Humans; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G / Q000097:blood*; D007129:Immunoglobulin Gm Allotypes
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D000939:Epitopes; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D007129:Immunoglobulin Gm Allotypes