The 512 monoclonal antibody specifically recognizes a 65-kDa and 55-kDa molecules of the erythrocyte membrane, which are the rat counterpart of mouse Crry/p65, a membrane inhibitor of complement component C3 convertase. Crry/p65 is widely distributed, predominantly expressed on endothelial cells and all circulating cells. This molecule was also detected on immature hepatocytes, systemic endothelial cells, skin fibroblasts, bronchial epithelial cells, bile canaliculi, the Schwann sheath of peripheral nerve fibers, and ependymal cells. Mouse Crry/p65 uses the same mechanisms as human Decay-Accelerating Factor (DAF) and human Membrane Cofactor Protein (MCP) in inhibiting C3 and C5 convertases, but it is not homologous to either DAF or MCP. It has been proposed that Crry/p65 is the mouse genetic homologue of human CR1 (CD35).
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.