CD223 Rat anti-Mouse, Brilliant Violet 650, Clone: C9B7W, BD Optibuild™

Description

• The C9B7W antibody monoclonal antibody specifically binds to an epitope in the D2 domain of CD223 (LAG3), the 70-kDa protein encoded by L ymphocyte- a ctivation g ene )
• A fusion protein consisting of the entire extracellular region of mouse LAG3 with mouse IgG1 was used as immunogen
• CD223 is a type-I membrane protein with four extracellular Ig-like domains; it is structurally homologous to CD4; and, like CD4, it binds MHC class II molecules
• However, unlike CD4, it is not expressed on resting human and mouse T lymphocytes
• In the mouse, as previously described in the human, CD223 expression is upregulated on T lymphocytes (both CD4+ and CD8+) activated through the T-cell receptor (TCR) and on IL-2-activated NK (LAK) cells, and it is not detected on B cells, dendritic cells, or Phorbol 12-myristate 13-acetate (PMA)-stimulated splenocytes
• Studies on human peripheral T lymphocytes suggest that CD223 associates with the TCR to downregulate TCR signaling
• In contrast, in vivo and in vitro evaluations of vaccination protocols in mice suggest that CD223 promotes immune responses by activating antigen-presenting cells
• Furthermore, NK cells of Lag3-/- mice display defects in their capacity to kill certain tumor cells
• Mouse CD223 also has been demonstrated to contribute to the suppressor function of T regulatory cells and the C9B7W antibody has been shown to inhibit this function in vitro and in vivo
• Therefore, CD223 appears to play complex roles in the regulation of immune responses
• Although the C9B7W antibody is unable to block the binding of MHC class II-IgG2a fusion protein to CD223, it is able to block the CD223-mediated inhibition of IL-2 production by a T-cell hybridoma responding to antigen
• The antibody was conjugated to BD Horizon™ BV650 which is part of the BD Horizon Brilliant™ Violet family of dyes
• This dye is a tandem fluorochrome of BD Horizon BV421 with an Ex Max of 405-nm and an acceptor dye with an Em Max at 650-nm
• BD Horizon BV650 can be excited by the violet laser and detected in a filter used to detect APC-like dyes (eg, 660/20-nm filter)
• Due to the excitation and emission characteristics of the acceptor dye, there will be spillover into the APC and Alexa Fluor™ 700 detectors
• However, the spillover can be corrected through compensation as with any other dye combination.