Fc Receptor Binding Asaay

Seattle Genova offers comprehensive Fc receptor & C1q binding tests with SPR technology. 

Fc Receptors

The human and non-human primate FcγR family consists of several activating receptors (FcγRI, FcγRIIa, FcγRIIIa) and a single inhibitory receptor, FcγRIIb. The complexity in Fc-function arises from differences in the specificity of human FcγRs and their affinities for IgG subclasses and can be further affected by genetic polymorphisms of the receptors.

There is one high-affinity IgG receptor in humans, hFcγRI (CD64), and 2 families of low-affinity IgG receptors, hFcγRIIA, IIB, and IIC (CD32), and hFcγRIIIA and IIIB (CD16). hFcγRI and hFcγRIIIA are FcRγ-associated activating receptors, hFcγRIIA and hFcγRIIC are single-chain activating receptors, hFcγRIIB are single-chain inhibitory receptors, and hFcγRIIIB are GPI-anchored receptors whose function is uncertain. The multiplicity of hFcγRs is further increased by a series of polymorphisms in their extracellular domains. 

IgG subclass binding profiles

C1q

C1q serves as a recognition and regulatory protein for the complement cascade. It is a protein complex involved in the complement system, which is part of the innate immune system. C1q together with C1r and C1s form the C1 complex. Antibodies of the adaptive immune system can bind antigen, forming an antigen-antibody complex. When C1q binds antigen-antibody complexes, the C1 complex becomes activated. Activation of the C1 complex initiates the classical complement pathway of the complement system. The antibodies IgM and all IgG subclasses except IgG4 can initiate the complement system.

Alternative Receptors for IgG

FcRL

Fc receptor-like proteins, consisting of six members (FCRL1-6) were originally identified as homologs of FcγR but were for a long time regarded as orphan receptors, mostly expressed on B-cells.

TRIM21

Tripartite motif-containing protein 21 (TRIM21) is a cytosolic protein expressed in almost all cell types but highly expressed in immune cells. TRIM21 was found to bind IgG with nanomolar affinity. TRIM21 binds IgG in the Fc domain at the CH2–CH3 interface similar to FcRn and protein A/G, it competes for binding to IgG with protein A/G, and binding is independent of N-glycosylation of the CH2 domain.

DC-SIGN

It has been hypothesized that DC-SIGN binds to the CH2–CH3 interface of the Fc domain of IgG, owing to the opening up of the site, where DC-SIGN binds due to the charged sialic acid.

Our services

The Fc receptor binding panel includes 10 individual Fc receptor assays, while the C1q binding test includes only C1q. For each receptor binding assay, the receptors are either immobilized directly on the chip or they are captured with an anti-his antibody is immobilized onto a CM5 chip. Multiple concentrations of testing protein flow through the Fc receptor. SPR sensorgrams are fit to obtain the affinity (KD) between the testing protein and Fc receptor.