Validated using a knockout cell line
Recombinant
RabMAb

Anti-MLH1 antibody [EPR3894] (ab92312)

Overview

  • Product name
    Anti-MLH1 antibody [EPR3894]
    See all MLH1 primary antibodies
  • Description
    Rabbit monoclonal [EPR3894] to MLH1
  • Host species
    Rabbit
  • Specificity
    The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.
  • Tested applications
    Suitable for: WB, IHC-P, Flow Cyt, ICC/IFmore details
    Unsuitable for: IP
  • Species reactivity
    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to Human MLH1 aa 700-800.
    Database link: P40692

  • Positive control
    • WB: 293, HeLa, HCT116, A431, and SW480 cell lysates; Human testis , Rat testis, Mouse thymus tissue lysates IHC-P: Human colonic adenocarcinoma and tonsil tissues. ICC/IF: HeLa and SW480 cells.
  • General notes

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMab® patents

    We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab92312 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
WB 1/2000. Predicted molecular weight: 84 kDa.

For unpurifid use at 1/10000 - 1/50000

IHC-P 1/100 - 1/250. Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol. PubMed: 22022465

Performed after boiling twice before IHC protocol carried out.

See IHC antigen retrieval protocols.

The mouse and rat recommendation is based on the WB results. We do not guarantee IHC-P for mouse and rat.

Flow Cyt 1/10 - 1/100.

ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.

 

ICC/IF 1/500.
  • Application notes
    Is unsuitable for IP.
  • Target

    • Function
      Heterodimerizes with PMS2 to form MutL alpha, a component of the post-replicative DNA mismatch repair system (MMR). DNA repair is initiated by MutS alpha (MSH2-MSH6) or MutS beta (MSH2-MSH6) binding to a dsDNA mismatch, then MutL alpha is recruited to the heteroduplex. Assembly of the MutL-MutS-heteroduplex ternary complex in presence of RFC and PCNA is sufficient to activate endonuclease activity of PMS2. It introduces single-strand breaks near the mismatch and thus generates new entry points for the exonuclease EXO1 to degrade the strand containing the mismatch. DNA methylation would prevent cleavage and therefore assure that only the newly mutated DNA strand is going to be corrected. MutL alpha (MLH1-PMS2) interacts physically with the clamp loader subunits of DNA polymerase III, suggesting that it may play a role to recruit the DNA polymerase III to the site of the MMR. Also implicated in DNA damage signaling, a process which induces cell cycle arrest and can lead to apoptosis in case of major DNA damages. Heterodimerizes with MLH3 to form MutL gamma which plays a role in meiosis.
    • Tissue specificity
      Colon, lymphocytes, breast, lung, spleen, testis, prostate, thyroid, gall bladder and heart.
    • Involvement in disease
      Defects in MLH1 are the cause of hereditary non-polyposis colorectal cancer type 2 (HNPCC2) [MIM:609310]. Mutations in more than one gene locus can be involved alone or in combination in the production of the HNPCC phenotype (also called Lynch syndrome). Most families with clinically recognized HNPCC have mutations in either MLH1 or MSH2 genes. HNPCC is an autosomal, dominantly inherited disease associated with marked increase in cancer susceptibility. It is characterized by a familial predisposition to early onset colorectal carcinoma (CRC) and extra-colonic cancers of the gastrointestinal, urological and female reproductive tracts. HNPCC is reported to be the most common form of inherited colorectal cancer in the Western world, and accounts for 15% of all colon cancers. Cancers in HNPCC originate within benign neoplastic polyps termed adenomas. Clinically, HNPCC is often divided into two subgroups. Type I: hereditary predisposition to colorectal cancer, a young age of onset, and carcinoma observed in the proximal colon. Type II: patients have an increased risk for cancers in certain tissues such as the uterus, ovary, breast, stomach, small intestine, skin, and larynx in addition to the colon. Diagnosis of classical HNPCC is based on the Amsterdam criteria: 3 or more relatives affected by colorectal cancer, one a first degree relative of the other two; 2 or more generation affected; 1 or more colorectal cancers presenting before 50 years of age; exclusion of hereditary polyposis syndromes. The term 'suspected HNPCC' or 'incomplete HNPCC' can be used to describe families who do not or only partially fulfill the Amsterdam criteria, but in whom a genetic basis for colon cancer is strongly suspected.
      Defects in MLH1 are a cause of mismatch repair cancer syndrome (MMRCS) [MIM:276300]; also known as Turcot syndrome or brain tumor-polyposis syndrome 1 (BTPS1). MMRCS is an autosomal dominant disorder characterized by malignant tumors of the brain associated with multiple colorectal adenomas. Skin features include sebaceous cysts, hyperpigmented and cafe au lait spots.
      Defects in MLH1 are a cause of Muir-Torre syndrome (MuToS) [MIM:158320]; also abbreviated MTS. MuToS is a rare autosomal dominant disorder characterized by sebaceous neoplasms and visceral malignancy.
      Note=Defects in MLH1 may contribute to lobular carcinoma in situ (LCIS), a non-invasive neoplastic disease of the breast.
      Defects in MLH1 are a cause of susceptibility to endometrial cancer (ENDMC) [MIM:608089].
      Note=Some epigenetic changes can be transmitted unchanged through the germline (termed 'epigenetic inheritance'). Evidence that this mechanism occurs in humans is provided by the identification of individuals in whom 1 allele of the MLH1 gene is epigenetically silenced throughout the soma (implying a germline event). These individuals are affected by HNPCC but does not have identifiable mutations in MLH1, even though it is silenced, which demonstrates that an epimutation can phenocopy a genetic disease.
    • Sequence similarities
      Belongs to the DNA mismatch repair mutL/hexB family.
    • Cellular localization
      Nucleus.
    • Information by UniProt
    • Database links
    • Alternative names
      • COCA 2 antibody
      • COCA2 antibody
      • DNA mismatch repair protein Mlh1 antibody
      • FCC 2 antibody
      • FCC2 antibody
      • hMLH 1 antibody
      • hMLH1 antibody
      • HNPCC 2 antibody
      • HNPCC antibody
      • HNPCC2 antibody
      • MGC5172 antibody
      • MLH 1 antibody
      • MLH1 antibody
      • MLH1_HUMAN antibody
      • MutL homolog 1 (E. coli) antibody
      • MutL homolog 1 antibody
      • MutL homolog 1 colon cancer nonpolyposis type 2 antibody
      • MutL homolog 1, colon cancer, nonpolyposis type 2 (E. coli) antibody
      • MutL protein homolog 1 antibody
      • MutL, E. coli, homolog of, 1 antibody
      see all

    Images

    • Immunocytochemistry/ Immunofluorescence analysis of SW480 (Human colorectal adenocarcinoma epithelial cell) cells labeling MLH1 with Purified ab92312 at 1:500 dilution (1.6 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor®594) 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor®488, ab150077) was used as the secondary antibody at 1:1000 (2 µg/ml) dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.
    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human colon tissue sections labeling MLH1 with Purified ab92312 at 1:250 dilution (2.9 µg/ml). Heat mediated antigen retrieval was performed using Perform heat mediated antigen retrieval using citrate (pH 6.0)ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody.Negative control:PBS instead of the primary antibody.Hematoxylinwas used as a counterstain
    • Immunocytochemistry/Immunofluorescence analysis of HeLa (human cervix adenocarcinoma) labelling MLH1 with purified ab92312 at 1/1000. Cells were fixed with 4% PFA and permeabilized with 0.1% Triton X-100. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody (Ab150077). Nuclei counterstained with DAPI (blue).

      Control: PBS only

    • Lane 1: Wild-type HAP1 cell lysate (20 µg)
      Lane 2: MLH1 knockout HAP1 cell lysate (20 µg)
      Lane 3: HCT116 cell lysate (20 µg)
      Lane 4: 293T cell lysate (20 µg)
      Lanes 1 - 4: Merged signal (red and green). Green - ab92312 observed at 88 kDa. Red - loading control, ab8245, observed at 37 kDa.


      Unpurified ab92312 was shown to recognize MLH1 in wild-type HAP1 cells along with additonal cross reactive bands. No band was observed when MLH1 knockout samples were examined. Wild-type and MLH1 knockout samples were subjected to SDS-PAGE. ab92312 and ab8245 (loading control to GAPDH) were both diluted 1/1000 and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1 hour at room temperature before imaging.

    • All lanes : Anti-MLH1 antibody [EPR3894] (ab92312) at 1/10000 dilution (Unpurified)

      Lane 1 : 293 cell lysate
      Lane 2 : HeLa cell lysate
      Lane 3 : A431 cell lysate
      Lane 4 : SW480 cell lysate

      Lysates/proteins at 10 µg per lane.

      Secondary
      All lanes : goat anti-rabbit HRP at 1/2000 dilution

      Predicted band size: 84 kDa

    • Unpurified ab92312 at 1/100 dilution staining MLH1 in Human colonic adenocarcinoma by Immunohistochemistry, Paraffin-embedded tissue. The use of an HRP/AP polymerized antibody is recommended for a secondary antibody.

    • Unpurified ab92312 (1/200) staining MLH1 in HeLa cells (green). Cells were fixed in paraformaldehyde, permeabilised with 0.5% Triton X100/PBS and counterstained with DAPI in order to highlight the nucleus (red). For further experimental details please refer to abreview.

      See Abreview

    • Overlay histogram showing HeLa cells stained with unpurified ab92312 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab92312, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.

    • Unpurified ab92312 staining MLH1 in Human colorectal (top) and gastric tissue (bottom) by Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections).

    • Unpurified ab92312 at 1/100 dilution staining MLH1 in Human tonsil by Immunohistochemistry, Paraffin-embedded tissue. The use of an HRP/AP polymerized antibody is recommended for a secondary antibody.

    References

    This product has been referenced in:
    • Shi L  et al. DNA methylation-mediated repression of miR-181a/135a/302c expression promotes the microsatellite-unstable colorectal cancer development and 5-FU resistance via targeting PLAG1. J Genet Genomics 45:205-214 (2018). WB . Read more (PubMed: 29735329) »
    • Pussila M  et al. Mlh1 deficiency in normal mouse colon mucosa associates with chromosomally unstable colon cancer. Carcinogenesis 39:788-797 (2018). Read more (PubMed: 29701748) »

    See all 17 Publications for this product

    Customer reviews and Q&As

    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (HeLa)
    Gel Running Conditions
    Reduced Denaturing (4-12%)
    Loading amount
    30 µg
    Specification
    HeLa
    Blocking step
    Milk as blocking agent for 45 minute(s) · Concentration: 2% · Temperature: 37°C
    Username

    Abcam user community

    Verified customer

    Submitted Jul 13 2015

    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 25°C
    Antigen retrieval step
    Heat mediated
    Sample
    Human Tissue sections (endometrial cancer)
    Specification
    endometrial cancer
    Permeabilization
    No
    Fixative
    Formaldehyde
    Username

    Abcam user community

    Verified customer

    Submitted Jul 23 2013

    Merci de confirmer ces détails et de votre coopération. Les détails fournis nous permettent de suivre de près la qualité de nos produits.

    Je suis désolé que ces produits ne fonctionnent pas co...

    Read More

    Merci beaucoup pour toutes ces informations détaillées qui nous sont très utiles pour identifier l'origine des problèmes rencontrés avec ab92312 et ab126057.

    Je n'ai rien à modifier à votre prot...

    Read More

    Je suis désolée d'apprendre que les anticorps ab92312 et ab126057 ne vous donnent pas de bons résultats en WB.

    Pour nous permettre de vous fournir la meilleure assistance technique, pourriez-vous nous donner les informatio...

    Read More
    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application
    Immunocytochemistry/ Immunofluorescence
    Sample
    Human Cell (HeLa)
    Specification
    HeLa
    Fixative
    Paraformaldehyde
    Permeabilization
    Yes - 0.5% Triton-X100 in PBS
    Username

    Dr. Kirk Mcmanus

    Verified customer

    Submitted Apr 18 2012

    Thank you for contacting us.
    I can confirm that the immunogen sequence of ab92312 is 100% similar with chicken MLH1 (http://www.uniprot.org/uniprot/E1BQE0)
    ab62729; the sequence is 81% similar to chicken MLH1 (http://www.uniprot.org/uniprot/E...

    Read More
    Abcam guarantees this product to work in the species/application used in this Abreview.
    Application
    Western blot
    Sample
    Human Cell lysate - whole cell (Various tumor cell lines)
    Loading amount
    30 µg
    Specification
    Various tumor cell lines
    Gel Running Conditions
    Reduced Denaturing
    Blocking step
    Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
    Username

    Abcam user community

    Verified customer

    Submitted Nov 03 2010

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

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