The 2ST8.5H7 monoclonal antibody specifically recognizes an epitope formed by the combination of CD8 alpha and beta chains. The majority of peripheral blood CD8+ T lymphocytes expresses a CD8αβ heterodimer (32, 30 kilodaltons (kDa)), while CD8+CD16+ natural killer (NK) cells and CD8+ TCR γδ+ T lymphocytes express CD8αα homodimers. The 2ST8.5H7 antibody can therefore be used to selectively bind to CD8+ T cells while excluding CD8+ NK cells. CD8 binds to class I major histocompatibility (MHC) molecules, resulting in increased adhesion between the CD8+ T lymphocytes and target cells. Binding of CD8 to class I MHC molecules enhances the activation of resting T lymphocytes. CD8 is coupled to a protein tyrosine kinase, p56lck. The CD8:p56lck complex can play a role in T-lymphocyte activation through mediation of the interactions between CD8 and the CD3/TCR complex. The CD8β antigen is present on the human suppressor/cytotoxic T-lymphocyte subset. The CD8 antigen is expressed on 19% to 48% of normal peripheral blood lymphocytes and 60% to 85% of normal thymocytes. The 2ST8.5H7 antibody crossreacts with lymphocytes of some nonhuman primate species.
The antibody was conjugated to BD Horizon™ BUV496 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 496-nm. BD Horizon BUV496 can be excited by the ultraviolet laser (355 nm) and detected with a 515/30 nm filter with a 450LP. Due to the excitation of the acceptor dye by other laser lines, there may be significant spillover into the channel detecting BD Horizon V500 or BV510 (eg, 525/40-nm filter). However, the spillover can be corrected through compensation as with any other dye combination.