Anti-H-2K Antibody, clone Y-3

41.30/39.15 kDa (pro-/mature H2-Kb alpha chain; UniProt P01901) and 13.78/11.64 kDa (pro-/mature β2-microglobulin; UniProt P01887) calculated. Note that the sequences vary among haplotypes/polymorphic forms and the actual observed molecular sizes will appear larger than the calculated values due to glycosylation.

Major histocompatibility complexes (MHCs) are cell surface molecules that play a major part of the immune system in all vertebrates by determining histocompatibility. The main function of MHCs is to bind and present peptide fragments derived from pathogens on the surface of antigen presenting cells. MHCs are divided into two major classes, class I MHCs are heterodimers composed of a transmembrane alpha chain and a common extracellular beta subunit. The alpha chain contains three domains (alpha1, alpha2, and alpha3), with the alpha1 domain mediating interaction with the beta2 subunit. The alpha1 and alpha2 domains form the antigen-binding groove. Class II MHCs are heterodimers of two single transmembrane chains, alpha and beta, each contains two domains, with the N-terminal alpha1 and beta1 domain from each chain forming the antigen-binding groove. Murine class I MHC is further divided into the classical H-2D, H-2K and H-2L subclasses, and the non-classical H-2Q, H-2M and H-2T subclasses. MHC molecules are highly polymorphic and laboratory mouse strains are homozygous with respect to their respective MHC haplotypes. The haplotype of each H-2K, for example, is specified by a lower case letter (b, d, k, q, r, s) in superscript. MHC class I molecules are ligands for the antigen receptors of CD8+ T cells and NK cells. MHC class I antigenic peptides assembly takes place within the endoplasmic reticulum (ER) and are facilitated by the peptide loading complex (PLC) composed of multiple proteins, including transporter associated with antigen processing (TAP), Tapasin, the thiol oxidoreductase ERp57 and calreticulin (CRT).

BALB.B mouse spleen cells.

Affects Function: A representative lot specifically killed B-cells derived from C57BL/6J (H-2 haplotype b) mice, but not B-cells from BALB/c (H-2 haplotype d) mice in a mixed B-cell culture (Jone, B., and Janeway, C.A. Jr. (1981). Nature. 292(5823):547-549).

Affinity Binding Assay: A representative lot reacted with cell surface murine class I MHC H-2K haplotypes b, k, q, r, s, but not d, in affinity binding assays utilizing 125I-labelled antibody. Clone Y-3 exhibited similar affinity toward seven H-2Kb mutant strains as the wild-type strain (Hämmerling, G.J., et al. (1982). Proc. Natl. Acad. Sci. U. S. A. 79(15):4737-4741).

Flow Cytometry Analysis: A representative lot detected the surface expression of exogenously transfected H-2Kb Y84C mutant among TAP- and tapasin-deficient BL/6 fibroblasts, while little exogenously expressed wild-type H-2Kb was detected on the cell surface in the absence of either TAP or the TAP-binding protein tapasin (Wijeyesakere, S.J., et al. (2015). Proc. Natl. Acad. Sci. U. S. A. 112(41):E5608-5617).

Flow Cytometry Analysis: Clone Y-3 ascites fluid detected an induciton of cell surface H-2K immunoreactivity upon exogenous murine calreticulin (mCRT) expression in the CRT-deficient K42 mouse fibroblast line (Del Cid, N., et al. (2010). J. Biol. Chem. 285(7):4520-4535).

Immunocytochemistry Analysis: A representative lot detected the surface expression of exogenously transfected H-2Kb Y84C mutant among TAP- and tapasin-deficient BL/6 fibroblasts, while exogenously expressed wild-type H-2Kb remained intracelluar in the absence of either TAP or the TAP-binding protein tapasin (Wijeyesakere, S.J., et al. (2015). Proc. Natl. Acad. Sci. U. S. A. 112(41):E5608-5617).

Immunoprecipitation Analysis: A representative lot immunoprecipitated H2-K from a calreticulin-/CRT-deficient mouse embryonic fibroblast (MEF) line transfected to express murine CRT (Wijeyesakere, S.J., et al. (2015). Proc. Natl. Acad. Sci. U. S. A. 112(41):E5608-5617).

This mouse monoclonal Anti-H-2K Antibody, clone Y-3, Cat. No. MABF946 is validated for use in Affinity Binding Assay, Flow Cytometry, Function Assay, Immunocytochemistry, and Immunoprecipitation for the detection of mouse class I MHC H-2K.

Clone Y-3 was isolated from a BALB/c (H-2 haplotype d) mouse immunized with Con A-stimulated murine spleen cells from the BALB.B (H-2 haplotype b) strain and originally referred to as anti-H-2.5. Clone Y-3 is reported to recognize murine class I MHC subclass H-2K haplotypes b, k, q, r, s, but not d (Hämmerling, G.J., et al. (1982). Proc. Natl. Acad. Sci. U. S. A. 79(15):4737-4741; Jone, B., and Janeway, C.A. Jr. (1981). Nature. 292(5823):547-549).

Format: Purified

Purified mouse IgG2b in PBS without preservatives.

Evaluated by Flow Cytometry in C57BL/6 mouse splenocytes.

Flow Cytometry Analysis: 0.1 µg of this antibody detected H-2K immunoreactivity on the surface of one million C57BL/6 mouse splenocytes (H-2 haplotype b).

Concentration: Please refer to lot specific datasheet.