Overview

  • Product name
    Anti-MHC class I antibody [ER-HR 52]
    See all MHC class I primary antibodies
  • Description
    Rat monoclonal [ER-HR 52] to MHC class I
  • Host species
    Rat
  • Specificity
    Ab15681 detects MHC class I antigens of d and b haplotypes.
  • Tested applications
    Suitable for: IHC-Fr, IHC (PFA fixed)more details
    Unsuitable for: IHC-FoFr
  • Species reactivity
    Reacts with: Mouse
  • Immunogen

    Murine macrophage precursor cells.

  • Positive control
    • IHC-Fr: Mouse spleen and brain tissue.
  • General notes


    Ab15681 detects MHC class I antigens and is therefore a valuable tool for studying cytotoxic T-cell interactions with class I positive antigen presenting cells.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Upon delivery aliquot. Avoid freeze / thaw cycle.
  • Storage buffer
    Preservative: 0.01% Thimerosal (merthiolate)
    Constituents: PBS, 10mg/ml BSA. pH 7.2
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Primary antibody notes
    Ab15681 detects MHC class I antigens and is therefore a valuable tool for studying cytotoxic T-cell interactions with class I positive antigen presenting cells.
  • Clonality
    Monoclonal
  • Clone number
    ER-HR 52
  • Isotype
    IgG2a
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab15681 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
IHC-Fr Use a concentration of 1 µg/ml. The determinant recognised by ab15681 is glutaraldehyde (0.05%), paraformaldehyde (1%) and acetone-resistant.
IHC (PFA fixed) 1/300. PubMed: 17446933
  • Application notes
    Is unsuitable for IHC-FoFr.
  • Target

    • Relevance
      MHC Class I molecules play a central role in the immune system by presenting peptides derived from the endoplasmic reticulum lumen. MHC class I antigens are heterodimers consisting of one alpha chain (44kDa) with beta 2 microglobulin (11.5 kDa). The antigen is expressed by all somatic cells at varying levels. MHC Class I molecules are expressed on most nucleated cells where they present endogenously synthesized antigenic peptides to CD8+ T lymphocytes, which are usually cytotoxic T cells. Fibroblasts or neurons however only show a low level of antigen.
    • Cellular localization
      Cell Membrane; Type I membrane protein.
    • Alternative names
      • A 28 antibody
      • A 9 antibody
      • Antigen presenting molecule antibody
      • Aw 24 antibody
      • Aw 68 antibody
      • CLASS I HISTOCOMPATIBILITY ANTIGEN antibody
      • H2 K1 antibody
      • H2K antibody
      • HLA A antibody
      • HLA class I histocompatibility antigen A 1 alpha chain antibody
      • HLA class I histocompatibility antigen A 11 alpha chain antibody
      • HLA class I histocompatibility antigen A 2 alpha chain antibody
      • HLA class I histocompatibility antigen A 24 alpha chain antibody
      • HLA class I histocompatibility antigen A 3 alpha chain antibody
      • HLA class I histocompatibility antigen A 30 alpha chain antibody
      • HLA class I histocompatibility antigen A 32 alpha chain antibody
      • HLA class I histocompatibility antigen A 68 alpha chain antibody
      • HLAA antibody
      • Leukocyte antigen class I A antibody
      • Major histocompatibility complex class I A antibody
      • MHC class I antigen A 1 antibody
      • MHC class I antigen A*11 antibody
      • MHC class I antigen A*2 antibody
      • MHC class I antigen A*24 antibody
      • MHC class I antigen A*3 antibody
      • MHC class I antigen A*30 antibody
      • MHC class I antigen A*32 antibody
      • MHC class I antigen A*68 antibody
      • MHC class I antigen HLA A heavy chain antibody
      • MHC class I heavy chain H2 K antibody
      see all

    Images

    • Frozen section of mouse spleen tissue stained for MHC class I using ab15681 at 1/200 dilution in immunohistochemical analysis. 

    • Immunohistochemical analysis of frozen mouse brain tissue, post-infection with Salmonella typhimurium, staining MHC Class I with ab15681.
      Tissue was blocked with 2% BSA, 10% normal rabbit serum, before incubating with primary antibody overnight. A biotinylated anti-rat IgG was used as the secondary antibody. Staining was detected using DAB.

    References

    This product has been referenced in:
    • Saranchova I  et al. Discovery of a Metastatic Immune Escape Mechanism Initiated by the Loss of Expression of the Tumour Biomarker Interleukin-33. Sci Rep 6:30555 (2016). IHC-Fr . Read more (PubMed: 27619158) »
    • Placke T  et al. Platelet-derived MHC class I confers a pseudonormal phenotype to cancer cells that subverts the antitumor reactivity of natural killer immune cells. Cancer Res 72:440-8 (2012). ICC/IF . Read more (PubMed: 22127925) »
    See all 7 Publications for this product

    Customer reviews and Q&As

    1-3 of 3 Q&A

    Answer

    Can you please tell me what kind of tissue you are trying to stain (species and organ), how long it was fixed in formalin, and if you included an antigen retrievals step before incubation with the antibody?

    The application that is listed on the datasheet, IHC (PFA-fixed) was added based on a reference listed on the datasheet,

    Erlebacher A et al. Constraints in antigen presentation severely restrict T cell recognition of the allogeneic fetus. J Clin Invest 117:1399-411 (2007).

    For staining with ab15681, they used this protocol:

    For MHC class I staining, tissues were fixed overnight at 4°C in 4% paraformaldehyde, equilibrated in 30% sucrose at 4°C, embedded in OCT compound (Tissue-Tek; Sakura Finetek), and sectioned at 5 μm. Sections were immersed in acetone for 5 minutes at room temperature, air-dried for 10 minutes, treated with 3% H2O2 in methanol for 20 minutes, then blocked in 1% BSA/3% goat serum/0.4% Triton X-100. Sections were then incubated overnight at 4°C with a 1:300 dilution of rat anti-mouse MHC class I (clone ER-HR52; Abcam), followed by biotinylated goat anti-rat antibodies (1:400 dilution; Jackson ImmunoResearch Laboratories), HRP-conjugated streptavidin, and biotin-tyramide amplification. Streptavidin–Alexa Fluor 594 was used as the final fluorochrome.

    http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1849983/

    Note that their samples are frozen, not paraffin-embedded, and that they detect using a tyramide amplification at the end of the protocol. What do you use after the biotinylated secondary antibody?

    Read More

    Answer

    My apologies for the late reply, I was away last week. I have received the following information from the source of ab15681: 1. Blocking endogenous biotin is important with the detection system used (biotinylated secondary). This may be improved. 2. Routinely fixed (4% PFA) tissue may be too much chemically altered - try 1% PFA or 0.05% glutaraldehyde to be more gentle. 3. The choice of secondary antibodies and corresponding blocking serum can have unexpected results - even with reagents from renowned companies. Try secondary reagents that are differently adsorbed (e.g. preadsorbed or adsorbed with serum from a different source), or try secondary from a different company. 4. We can offer a refund if everything else fails. I hope this advice will help, please let us know if you still have a problem and we can arrange a credit note or refund,

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    Answer

    I'm very sorry to hear you are having a problem with ab15681. This antibody has not been tested in IHC on perfusion formaldehyde fixed sections, we recommend using fresh frozen sections followed by postfixation in glutaraldehyde (0.05%), paraformaldehyde (1%) or ice cold acetone. I will enquire with the originator of the antibody if they can suggest modifications to your protocol and a good positive control (lymphocytes are highly positive whereas fibroblasts or neurons show only a low level of antigen). My apologies for the delay,

    Read More

    Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

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