The CG4.rMab is a recombinant monoclonal antibody that was derived from CG4 hybridoma cells. The CG4.rMab specifically binds to human G-protein coupled receptor 56 (GPR56) like the conventional CG4 antibody and performs like the CG4 antibody when used to stain cells and analyze them by flow cytometry. GPR56 is also known as adhesion G-protein coupled receptor G1 (ADGRG1), or TM7XN1. GPR56 is a ~15kDa G protein-coupled receptor encoded by ADGRG1 which belongs to the adhesion-GPCR family that comprises 33 members in human. The extracellular region contains a mucin-like domain followed by a membrane proximal GPCR-autoproteolysis inducing (GAIN) domain, seven transmembrane regions and a cytoplasmic tail. The constitutive self-cleavage at the proteolytic site gives rise to a membrane spanning (C-terminal fragment or CTF) and an extracellular (N-terminal fragment or NTF) subunit that remain noncovalently bound, leading to the expression of a heterodimeric receptor at the cell surface. GPR56 is widely expressed with the highest levels of messenger found in the brain, heart, and thyroid gland. Recently, GPR56 was found to be variably expressed on platelets, cytotoxic NK cells and T lymphocytes including CD4+, CD8+, and γδ T cells. It was shown that GPR56 functions as an inhibitory receptor on NK cells through interaction with CD81. While GPR56 NTF associates with Tissue transglutaminase 2 and Collagen III (α-1), GPR56 CTF can recruit Gα proteins leading to the activation of mTOR and RhoA signaling pathways. GPR56 has been implicated in cell-cell interactions, adhesion, migration, and regulation of cell proliferation and survival of various cell types. New evidence also shows a role of GPR56 in tumor progression. Recently, the CG4 antibody was found to activate GPR56 in melanoma cells leading to an increase of IL-6 secretion, in a CD9/CD81-dependent manner.
The antibody was conjugated to BD Horizon BUV737 which is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This dye is a tandem fluorochrome with an Ex Max near 350 nm and an Em Max near 737 nm. BD Horizon Brilliant BUV737 can be excited by the ultraviolet laser (355 nm) and detected with a 740/35 nm filter. Due to the excitation of the acceptor dye by the red laser line, there may be significant spillover into red laser detectors with filters in the 700-720 nm range.