The ITEM-1 monoclonal antibody specifically binds to CD266, which is also known as TWEAK Receptor (TWEAK-R/TWEAKR), or Fibroblast growth factor inducible 14 (FGF-inducible 14, FN14). CD266 is a ~14 kDa type I transmembrane protein that is the 12A family member of the tumor necrosis factor receptor superfamily (TNFRSF12A). It is expressed at low levels in a variety of tissues including, heart, placenta, lung, muscle, and pancreas. CD266 is relatively highly expressed on HUVEC cells and certain tumor cell lines. TWEAK-induced CD266-mediated signaling can play roles in inflammation, the induction of apoptosis in certain cell types, the proliferation and migration of endothelial cells, and can promote angiogenesis within healthy and diseased tissues, eg, tumors. The ITEM-1 antibody can reportedly induce the proliferation of HUVEC cells and the death of certain TWEAK-sensitive tumor target cell lines. CD266 binds TWEAK (CD255) and can signal intracellularly by interactions with TRAF1, 2, and 3 cytoplasmic proteins leading to NF-κB activation.
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.