The P-3E10 monoclonal antibody specifically recognizes CD298, the β3 subunit of the Na+/K+ ATPase (Sodium/potassium-transporting ATPase subunit beta-3). CD298 is a ~45-55 kDa single-pass, type II membrane protein that is encoded by ATP1B3 (ATPase Na+/K+ transporting subunit beta 3) which belongs to the P-type ATPases superfamily. CD298 is widely expressed on lymphocytes, monocytes, granulocytes, platelets, and on other hematopoietic and nonhematopoietic cells and cell lines. The Na+/K+ ATPase is an integral membrane protein complex with enzymatic activity that mediates the active transport and exchange of sodium and potassium ions across the plasma membrane. This complex is composed of α and β subunits. The α subunit is a 10-membrane-spanning, catalytic protein that contains binding sites for Na +, K + and ATP. The α subunits are associated with the smaller, regulatory glycoprotein β subunits. Upon ATP hydrolysis, the Na+/K+ ATPase transports Na+ ions out of the cell in exchange for K+ ions that are transported in. This establishes a transmembrane electrochemical gradient that is essential for osmoregulation and for the transport of various nutrients and other molecules by cells. The P-3E10 antibody can reportedly inhibit the activation and proliferation of T cells and 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.