The 5B11 monoclonal antibody specifically recognizes mouse CD123, the α chain subunit (IL-3Rα) of the mouse IL-3 receptor. The IL-3Rα chain is a 60-70 kD transmembrane glycoprotein that binds IL-3 with low affinity but cannot transduce signals by itself. The mouse IL-3Rα chain can complex with either of two homologous β chain subunits (βc and βIL-3) to form high-affinity heterodimeric IL-3 receptors. The βc chain is the common β chain subunit that can complex with the α subunits of the mouse IL-3R, IL-5R and GM-CSFR to form high-affinity receptors. The βIL-3 subunit can form high affinity IL-3 receptor complexes with only the IL-3Rα subunit. The βIL-3 subunit clone is specific for mouse IL-3 and binds with low affinity. The immunogen used to generate the 5B11 hybridoma was CTLL cells transfected with the mouse IL-3R alpha chain. The 5B11 antibody does not block the binding of IL-3 protein to the high affinity IL-3 receptor heterodimer.
The antibody was conjugated to BD Horizon™ BV650 which is part of the BD Horizon Brilliant™ Violet family of dyes. This dye is a tandem fluorochrome of BD Horizon BV421 with an Ex Max of 405-nm and an acceptor dye with an Em Max at 650-nm. BD Horizon BV650 can be excited by the violet laser and detected in a filter used to detect APC-like dyes (eg, 660/20-nm filter). Due to the excitation and emission characteristics of the acceptor dye, there will be spillover into the APC and Alexa Fluor® 700 detectors. However, the spillover can be corrected through compensation as with any other dye combination.