The L203.rMab is a recombinant monoclonal antibody that was derived from L203 hybridoma cells. The L203.rMab specifically recognizes a monomorphic epitope on the extracellular region of human HLA-DR antigens which are human Major Histocompatibility Complex (MHC) Class II antigens. HLA-DR antigens are heterodimers comprised of two different type I transmembrane glycoproteins that are noncovalently-associated. The ~34 kDa HLA-DR alpha (HLA-DRα) chain is encoded by HLA-DRA whereas the ~28 kDa HLA-DR beta (HLA-DRβ) chains are encoded by one of the 4 different HLA-DRB loci (HLA-DRB1,3,4,5) that are located within the Human Leukocyte Antigen (HLA) Complex of chromosome 6. Each chain is comprised of an extracellular region with an IgSF domain, followed by a transmembrane sequence and a short cytoplasmic tail. The L203.rMab antibody recognizes a common determinant that is dependent on the association of HLA-DR alpha and beta chains. HLA-DR is variably expressed on B cells, activated T cells and NK cells, monocytes, macrophages, dendritic cells (DC), Langerhans cells, thymic epithelial cells, and tumor cell lines including B cell lines, myelomas, and some myeloid leukemias. HLA-DR functions in the presentation of peptide antigens to CD4+ T lymphocytes in the generation and regulation of adaptive immune responses. HLA-DR expressed on thymic stromal cells plays a key role in the positive and negative selection of CD4+ T cells during thymopoiesis. Certain HLA-DR alleles, polymorphisms or aberrant expression patterns are associated with susceptibility to diseases including autoimmunity and cancer. L203 antibody binding is reportedly blocked by the L243 monoclonal antibody suggesting that the two antibodies recognize crossreactive or spatially related determinants on HLA-DR.
The antibody was conjugated to BD Horizon™ BUV563 which is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This dye is a tandem fluorochrome of BD Horizon BUV395 which has an Ex Max of 348 nm and an acceptor dye. The tandem has an Em Max at 563 nm. BD Horizon BUV563 can be excited by the 355 nm ultraviolet laser. On instruments with a 561 nm Yellow-Green laser, the recommended bandpass filter is 585/15 nm with a 535 nm long pass to minimize laser light leakage. When BD Horizon BUV563 is used with an instrument that does not have a 561 nm laser, a 560/40 nm filter with a 535 nm long pass may be more optimal. Due to the excitation and emission characteristics of the acceptor dye, there may be spillover into the PE and PE-CF594 detectors. However, the spillover can be corrected through compensation as with any other dye combination.