The ACK45 monoclonal antibody specifically binds to CD117 (c-Kit), a transmembrane tyrosine-kinase receptor which is encoded by the Kit gene (formerly dominant white spotting, W). The c-Kit ligand (also known as steel factor, stem cell factor, and mast-cell growth factor) encoded by the Kitl gene (formerly steel, Sl), is a co-mitogen for hematopoietic stem cells and the myeloerythroid progenitors and a mast-cell differentiation factor. The Kit[W] and Kitl[Sl] mutant alleles have similar pleiotropic effects on the development of melanocytes, germ cells, and the hematopoietic system. In the adult bone marrow, CD117 is expressed on hematopoietic progenitor cells, including the CD90 (Thy-1)[lo] TER-119[-] CD45R/B220[-] CD11b (Mac-1)[-] Ly-6G (Gr-1)[-] CD4[-] CD8[-] Sca-1 (Ly-6A/E)[+] multipotent hematopoietic stem cells, progenitors committed to myeloid and/or erythroid lineages, and precursors of B and T lymphocytes. This widespread expression of CD117 in hematopoietic precursors is consistent with the participation of c-Kit and its ligand in the regulation of several hematopoietic lineages. Intrathymic expression of c-Kit and c-Kit ligand suggest that CD117 is also involved in the regulation of some events during the development of T lymphocytes. CD117 is also expressed by mast cells. The expression of CD117 has been included in protocols to isolate hematopoietic progenitors from fetal and adult peripheral blood. The specificity and functional activity of mAb ACK45 is very similar to that of the published antibody ACK22; it is reported to block the action of c-Kit in vitro and in vivo.
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 of BD Horizon BUV395 with an Ex Max of 348-nm and an acceptor dye with an Em Max at 737-nm. BD Horizon Brilliant BUV737 can be excited by the ultraviolet laser (355 nm) and detected with a 740/35 filter. Due to the excitation of the acceptor dye by other laser lines, there may be significant spillover into channels detecting Alexa Fluor® 700-like dyes (eg, 712/20-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 BUV737 reagents. Therefore, the use of BD CompBeads or BD CompBeads Plus to determine spillover values for these reagents is not recommended. Different BUV737 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.