The WT.5 monoclonal antibody specifically recognizes the alpha subunit of LFA-1 (αLβ2 integrin, CD11a/CD18), a heterodimeric surface glycoprotein which is found on the majority of leukocytes, but not on peritoneal macrophages or peritoneal mast cells. LFA-1 mediates a variety of heterotypic and homotypic intercellular adhesions through interaction with ICAM-1 (CD54) and ICAM-2 (CD102). WT.1 mAb recognizes both the activated and unactivated forms of LFA-1. It inhibits the binding of LFA-1 to ICAM-1 in several in vitro assays, including binding of Concanavalin A-stimulated lymphocytes (Con A blasts) to purified ICAM-1 and Mg2+-dependent aggregation of concanavalin A-stimulated blasts. It has also been reported to inhibit leukocyte infiltration in several in vivo models of inflammation.
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.