Product nameAnti-MHC Class II antibody [NIMR-4]
See all MHC Class II primary antibodies
DescriptionRat monoclonal [NIMR-4] to MHC Class II
Specificityab25333 reacts specifically with a non-polymorphic I-A-encoded epitope on MHC Class II antigens.
Tested applicationsSuitable for: Flow Cyt, ICC/IF, IHC-P, Functional Studies, IHC-Frmore details
Species reactivityReacts with: Mouse
The details of the immunogen for this antibody are not available.
- ICC/IF: Raw264.7 cells.
Abcam is committed to meeting high quality standards of ethical manufacturing and has decided to discontinue this product by June 2020 as it has been generated by the ascites method. We are sorry for any inconvenience this may cause. We suggest ab23990 as a possible replacement.
Storage instructionsShipped at 4°C. Upon delivery aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
Storage bufferpH: 8.20
Constituent: 100% Borate buffered saline
Concentration information loading...
Purification notesPurified from ascites.
Light chain typekappa
Our Abpromise guarantee covers the use of ab25333 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Flow Cyt||Use 1µl for 106 cells.
ab18536 - Rat monoclonal IgG2b, is suitable for use as an isotype control with this antibody.
|ICC/IF||Use at an assay dependent concentration.|
|IHC-P||1/150. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.|
|Functional Studies||Use at an assay dependent concentration.
Complement-dependent cytotoxicity assays.
FunctionBinds peptides derived from antigens that access the endocytic route of antigen presenting cells (APC) and presents them on the cell surface for recognition by the CD4 T-cells. The peptide binding cleft accomodates peptides of 10-30 residues. The peptides presented by MHC class II molecules are generated mostly by degradation of proteins that access the endocytic route, where they are processed by lysosomal proteases and other hydrolases. Exogenous antigens that have been endocytosed by the APC are thus readily available for presentation via MHC II molecules, and for this reason this antigen presentation pathway is usually referred to as exogenous. As membrane proteins on their way to degradation in lysosomes as part of their normal turn-over are also contained in the endosomal/lysosomal compartments, exogenous antigens must compete with those derived from endogenous components. Autophagy is also a source of endogenous peptides, autophagosomes constitutively fuse with MHC class II loading compartments. In addition to APCs, other cells of the gastrointestinal tract, such as epithelial cells, express MHC class II molecules and CD74 and act as APCs, which is an unusual trait of the GI tract. To produce a MHC class II molecule that presents an antigen, three MHC class II molecules (heterodimers of an alpha and a beta chain) associate with a CD74 trimer in the ER to form an heterononamer. Soon after the entry of this complex into the endosomal/lysosomal system where antigen processing occurs, CD74 undergoes a sequential degradation by various proteases, including CTSS and CTSL, leaving a small fragment termed CLIP (class-II-associated invariant chain peptide). The removal of CLIP is facilitated by HLA-DM via direct binding to the alpha-beta-CLIP complex so that CLIP is released. HLA-DM stabilizes MHC class II molecules until primary high affinity antigenic peptides are bound. The MHC II molecule bound to a peptide is then transported to the cell membrane surface. In B cells, the interaction between HLA-DM and MHC class II molecules is regulated by HLA-DO. Primary dendritic cells (DCs) also to express HLA-DO. Lysosomal miroenvironment has been implicated in the regulation of antigen loading into MHC II molecules, increased acidification produces increased proteolysis and efficient peptide loading.
Sequence similaritiesBelongs to the MHC class II family.
Contains 1 Ig-like C1-type (immunoglobulin-like) domain.
Cellular localizationCell membrane. Endoplasmic reticulum membrane. Golgi apparatus > trans-Golgi network membrane. Endosome membrane. Lysosome membrane. The MHC class II complex transits through a number of intracellular compartments in the endocytic pathway until it reaches the cell membrane for antigen presentation.
- Information by UniProt
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ab25333 staining MHC Class II in mouse spleen, white pulp tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde, permeabilized with 0.1% Tween-20 in PBS and blocked with 1% BSA for 40 minutes at room temperature; antigen retrieval was by heat mediation in citrate buffer pH 6. Samples were incubated with primary antibody (1/150 in TBS + 1% BSA) for 18 hours at 4°C. A Biotin-conjugated goat anti-rat IgG polyclonal (1/100) was used as the secondary antibody.
Flow Cytometry analysis of BALB/c mouse splenocytes labeling MHC Class II with ab25333 at 1 μg/106 cells (purple). A Mouse Anti-Rat IgG2b-AF488 was used as the secondary antibody. Grey - Isotype Control, Rat IgG2b-UNLB, followed by Mouse Anti-Rat IgG2b-AF488.
ICC/IF image of ab25333 stained Raw264.7 cells. The cells were 4% Formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal Goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab25333, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab98420, DyLight® 488 Goat anti-Rat IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
Acetone fixed Mouse LLC tumour tissue stained for MHC Class II using ab25333 at 1/150 dilution in immunohistochemical analysis. The tissue was incubated for 18 hours at 4°C and blocked using BSA for 1 hour at room temperature. Alexa Fluor® 488 Goat Anti-Rat IgG (H+L) Antibody was used as the secondary at 1/100 dilution (green).
ab25333 has been referenced in 10 publications.
- Zhou B et al. Assisting anti-PD-1 antibody treatment with a liposomal system capable of recruiting immune cells. Nanoscale 11:7996-8011 (2019). PubMed: 30969294
- Hakim R et al. Mesenchymal stem cells transplanted into spinal cord injury adopt immune cell-like characteristics. Stem Cell Res Ther 10:115 (2019). PubMed: 30944028
- Lin X et al. Combined a-programmed death-1 monoclonal antibody blockade and fractionated radiation therapy reduces tumor growth in mouse EL4 lymphoma. Cancer Biol Ther 20:666-679 (2019). PubMed: 30572778
- Quaranta V et al. Macrophage-Derived Granulin Drives Resistance to Immune Checkpoint Inhibition in Metastatic Pancreatic Cancer. Cancer Res 78:4253-4269 (2018). PubMed: 29789416
- Choi SH et al. Tumour-vasculature development via endothelial-to-mesenchymal transition after radiotherapy controls CD44v6+ cancer cell and macrophage polarization. Nat Commun 9:5108 (2018). PubMed: 30504836
- Lee HJ et al. Glucocorticoids induce corneal allograft tolerance through expansion of monocytic myeloid-derived suppressor cells. Am J Transplant N/A:N/A (2018). PubMed: 30019411
- Liao T et al. In Vivo Attenuation of Antibody-Mediated Acute Renal Allograft Rejection byEx VivoTGF-ß-Induced CD4+Foxp3+Regulatory T Cells. Front Immunol 8:1334 (2017). PubMed: 29085374
- Vethanayagam RR et al. Toll-like receptor 4 modulates the cochlear immune response to acoustic injury. Cell Death Dis 7:e2245 (2016). PubMed: 27253409
- Yang W et al. Activation of the antigen presentation function of mononuclear phagocyte populations associated with the basilar membrane of the cochlea after acoustic overstimulation. Neuroscience 303:1-15 (2015). IHC ; Mouse . PubMed: 26102003
- Rocca CJ et al. Treatment of Inherited Eye Defects by Systemic Hematopoietic Stem Cell Transplantation. Invest Ophthalmol Vis Sci 56:7214-23 (2015). PubMed: 26540660