The 28-14-8 antibody reacts with the α3 domain of the H-2D[b] MHC class I alloantigen. The antibody binds to H-2D[b] in the presence or absence of the β2 microglobulin chain. It cross-reacts with the α3 domain of H-2L[d], but not with K[d] or D[d], and with H-2D[q] and/or L[q]. Reactivity with haplotypes k, f, p, r, and s has not been observed. mAb 28-14-8 has been shown to block H-2L[d]- specific and H- 2L[d]-restricted antigen-specific lysis of target cells by cytotoxic T lymphocytes (CTL), but it does not block recognition of H-2L[d] positive target cells by Ly-6G2-positive NK 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.