The G28-8 monoclonal antibody specifically recognizes RP105/Bgp95, a 95-105 kDa type I membrane protein consisting of extracellular leucine-rich repeats and a short cytoplasmic domain. It is expressed on mantle zone B cells, but weakly on germinal center B cells. RP105/Bgp95 is also expressed on peripheral blood monocytes, dendritic cells, and a subset of peripheral blood lymphocytes. The extracellular domain associates with a molecule called MD-1 to form a cell surface receptor complex RP105/Bgp95/MD- 1. This receptor belongs to the family of toll-like receptors (TLR). Studies show that RP105/Bgp95/MD-1, working in concert with TLR4, controls B cell recognition and signaling of lipopolysaccharide (LPS). Reports on functional studies show that G28-8 monoclonal antibody can induce a G0 to G1 cell cycle transition and was synergistic with PMA, anti-µ, or anti-CD40 in inducing proliferation of resting B cells.
The antibody was conjugated to BD Horizon™ BUV661 which is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This dye is a tandem fluorochrome of BD Horizon BUV395 with an Ex Max of 348-nm and an acceptor dye with an Em Max at 661-nm. BD Horizon Brilliant BUV661 can be excited by the ultraviolet laser (355 nm) and detected with a 670/25 filter and a 630 nm LP. Due to cross laser excitation of this dye, there may be significant spillover into channels detecting APC-like emissions (eg, 670/25-nm filter).
Due to spectral differences between labeled cells and beads, using BD™ CompBeads can result in incorrect spillover values when used with BD Horizon BUV661 reagents. Therefore, the use of BD CompBeads or BD CompBeads Plus to determine spillover values for these reagents is not recommended. Different BUV661 reagents (eg, CD4 vs. CD45) can have slightly different fluorescence spillover therefore, it may also be necessary to use clone-specific compensation controls when using these reagents.