Mouse anti Human HLA Class I Heavy Chain (Restricted expression)

The HLA class I gene family is composed of a group of genes whose products encode cell surface glycoproteins of MW 40–45 kDa, associated non-covalently with the beta-2-microglobulin light chain. They include the three polymorphic molecules HLA-A, -B, and -C, which are ubiquitously expressed and which are able to present intracellular peptides to cytotoxic T cells. Three additional class I genes are known, commonly referred to as non-classical or class Ib genes, all highly homologous to the other class I genes and all of which associate with beta-2-microglobulin light chain. In Humans, each of the class Ib genes appears to exhibit a distinct pattern of expression in developing and adult tissues. HLA-E transcripts are distributed widely in adult tissues and have also been found in the placenta and fetal liver. In the adult, the presence of HLA-F has been shown in skin, resting T cells, and B cells, whereas its expression during development has been reported in fetal liver and at low levels in placenta and extra-placental tissues. HLA-G was originally thought to be expressed only in certain populations of placental trophoblasts, but low levels have also been found in a variety of Human tissues. Recently it was shown that HLA class I expression in breast cancer cells can have a predictive value for chemotherapy response.

HCA2 is a mouse monoclonal IgG1 antibody derived by fusion of SP2/0-Ag14 mouse myeloma cells with spleen cells from BALB/c mice immunized with HLA-B7 and -B40 heavy chains.

The antibody HCA2 reacts preferentially with HLA-A locus heavy chains. HCA2 was raised against free class I heavy chains of HLA, to obtain antibodies that would still react with denatured class I antigens, as they occur in Western blotting, conventional light microscopical analysis of formalin-fixed, paraffin-embedded sections, and cryo-immuno-electron microscopy. HCA2 indeed retains strong reactivity with free class I heavy chains in Western blots. HCA2 in particular reacts in a locus-specific manner by biochemical criteria. Conditions are described for use of HCA2 in immunohistochemical staining of formalin-fixed, paraffin-embedded sections (see references). HCA2 also produces strong reactivity in immuno-electron microscopy. Its use allows the determination of tissue and subcellular distribution of class I antigens. Optimal antibody dilutions for the different applications should be determined by titration; recommended range is 1:100 – 1:200 for flow cytometry, and for immunohistochemistry with avidin-biotinylated horseradish peroxidase complex (ABC) as detection reagent, and 1:100 – 1:1000 for immunoblotting applications.

1. Stam NJ, Vroom TM, Peters PJ, Pastoors EB, Ploegh HL. HLA-A- and HLA-B-specific monoclonal antibodies reactive with free heavy chains in western blots, in formalin-fixed, paraffin-embedded tissue sections and in cryo-immuno-electron microscopy. Int. Immunol. 1990;2:113-125._x000D_ 2. de Kruijf EM, van Nes JGH, Sajet A, Tummers QRJG, Putter H, Osanto S, Speetjens FM, Smit VTHBM, Liefers GJ, van de Velde CJH, Kuppen PJK. The predictive value of HLA Class I tumor cell expression and presence of intratumoral Tregs for chemotherapy in patients with early breast cancer. Clin Cancer Res 2010;16:1272-1280._x000D_ 3. Hutter H, Hammer A, Blaschitz A, Hartmann M, Ebbesen P, Dohr G, Ziegler A, Uchanska-Ziegler B. Expression of HLA class I molecules in Human first trimester and term placenta trophoblast. Cell Tissue Res 1996; 286:439–447. _x000D_ 4. Seitz C, Uchanska-Ziegler B, Zank A, Ziegler A. The monoclonal antibody HCA2 recognises a broadly shared epitope on selected classical as well as several non-classical HLA class I molecules. Mol Immunol. 1998;35:819-27._x000D_ 5. Goodridge JP, Burian A, Lee N, Geraghty DE. HLA-F and MHC class I open conformers are ligands for NK cell Ig-like receptors. J Immunol. 2013;191(7):3553-62

The antibody is shipped at ambient temperature and may be stored at +4°C; For prolonged storage prepare appropriate aliquots and store at or below -20°C; Prior to use, an aliquot is thawed slowly in the dark at ambient temperature, spun down again and used to prepare working dilutions by adding sterile phosphate buffered saline (PBS, pH 7.2); Repeated thawing and freezing should be avoided; Working dilutions should be stored at +4°C, not refrozen, and preferably used the same day; If a slight precipitation occurs upon storage, this should be removed by centrifugation; It will not affect the performance or the concentration of the product