Highly specific inhibition of C1q globular-head binding to human IgG: a novel approach to control and regulate the classical complement pathway using an engineered single-chain antibody variable fragment.
Based on these studies and the crystallographic model of Protein A binding to human IgG, we tested the three previously reported sites that demonstrated the best cross-linking (F5, F13, Q32) and nine additional sites in Protein Z for BPA incorporation.
Particularly, FCGR2A and FCGR3A possess functional variants, including missense mutations (FCGR2A H131R and FCGR3A V176F) that decrease their binding affinity to human IgG and affect the clearance of immune complexes [ 8, 18, 21, 23- 25].
Briefly, AQP4-IgG and MOG-IgG positivity was determined by visualization of binding to human embryonic kidney cells, transfected with the extracellular and trans-membrane domains of MOG or with full-length M23-AQP4.
Lack of core fucose in the Fc glycan enhanced the binding of IgG to FcγRIIIa up to 50-fold, together with slightly improved binding to the Arg131 FcγRIIa polymorphic form and FcγRIIb whereas the absence of fucose did not affect the binding to human FcγRI, C1q, and neonatal FcR.
However, the monoclonal IgG antibodies used in the study could be considered as surrogates for human IgEs, based on their ability to inhibit the binding of human IgG to the target allergen.
The sensitivity was limited by nonspecific binding of human IgG to the solid phase.
We used human serum samples diluted 1 100 and peroxidase-conjugated goat anti-human IgG (Clinisciences, Nanterre, France) diluted 1 20,000 to detect binding of human IgG.
Background binding to an irrelevant human IgG, MonoD, was subtracted.
For detection of binding of human IgG subclasses to FcγRs, the plate was coated with soluble FcγRIIa or FcγRIIb or FcγRIIIa at a concentration 5 µg/ml for 20 hours at 4°C.
