Overview

  • Product name

    Anti-Cytokeratin 20 antibody [EPR1621(2)]
    See all Cytokeratin 20 primary antibodies
  • Description

    Rabbit monoclonal [EPR1621(2)] to Cytokeratin 20
  • Host species

    Rabbit
  • Tested applications

    Suitable for: Flow Cyt, ICC/IF, WB, IPmore details
    Unsuitable for: IHC-P
  • Species reactivity

    Reacts with: Mouse, Rat, Human
  • Immunogen

    Synthetic peptide corresponding to residues in Human Cytokeratin 20 (P35900).

  • Positive control

    • WB: SW480, Mouse small intestine and Rat small intestine lysates ICC/IF: SW480 cells
  • General notes

    A trial size is available to purchase for this antibody.

     

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

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab109111 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
Flow Cyt Use at an assay dependent concentration.
ICC/IF 1/50 - 1/100.
WB 1/10000 - 1/50000. Detects a band of approximately 46 kDa (predicted molecular weight: 48 kDa).
IP 1/10 - 1/100.
  • Application notes
    Is unsuitable for IHC-P.
  • Target

    • Function

      Plays a significant role in maintaining keratin filament organization in intestinal epithelia. When phosphorylated, plays a role in the secretion of mucin in the small intestine.
    • Tissue specificity

      Expressed predominantly in the intestinal epithelium. Expressed in luminal cells of colonic mucosa. Also expressed in the Merkel cells of keratinized oral mucosa; specifically at the tips of some rete ridges of the gingival mucosa, in the basal layer of the palatal mucosa and in the taste buds of lingual mucosa.
    • Sequence similarities

      Belongs to the intermediate filament family.
    • Developmental stage

      First detected at embryonic week 8 in individual 'converted' simple epithelial cells of the developing intestinal mucosa. In later fetal stages, synthesis extends over most goblet cells and a variable number of villus enterocytes. In the developing gastric and intestinal mucosa, expressed in all enterocytes and goblet cells as well as certain 'low-differentiated' columnar cells, whereas the neuroendocrine and Paneth cells are negative.
    • Post-translational
      modifications

      Hyperphosphorylation at Ser-13 occurs during the early stages of apoptosis but becomes less prominent during the later stages. Phosphorylation at Ser-13 also increases in response to stress brought on by cell injury.
      Proteolytically cleaved by caspases during apoptosis. Cleavage occurs at Asp-228.
    • Cellular localization

      Cytoplasm.
    • Information by UniProt
    • Database links

    • Alternative names

      • CD20 antibody
      • CK 20 antibody
      • CK-20 antibody
      • CK20 antibody
      • Cytokeratin-20 antibody
      • Cytokeratin20 antibody
      • K1C20_HUMAN antibody
      • K20 antibody
      • KA20 antibody
      • Keratin 20 antibody
      • keratin 20, type I antibody
      • keratin 21, rat, homolog of antibody
      • Keratin antibody
      • Keratin type I cytoskeletal 20 antibody
      • Keratin-20 antibody
      • Keratin20 antibody
      • KRT 20 antibody
      • KRT 21 antibody
      • KRT20 antibody
      • KRT21 antibody
      • MGC35423 antibody
      • OTTHUMP00000164518 antibody
      • Protein IT antibody
      • type I cytoskeletal 20 antibody
      see all

    Images

    • Flow Cytometry analysis of SW480 (human colorectal adenocarcinoma ) cells labeling Cytokeratin 20 with unpurified ab109111 at 1:20 dilution(10ug/ml) (red). Cells were fixed with 4% paraformaldehyde and permeabilised with 90% methanol. A Goat anti rabbit IgG (Alexa Fluor® 488)(ab150077)(1:2000 dilution) was used as the secondary antibody. Rabbit monoclonal IgG (Black)(ab172730) was used as the isotype control, Cell without incubation with primary antibody and secondary antibody (Blue) were used as the unlabeled control.

    • All lanes : Anti-Cytokeratin 20 antibody [EPR1621(2)] (ab109111) at 1/10000 dilution

      Lane 1 : SW480 lysate
      Lane 2 : Mouse small intestine lysate
      Lane 3 : Rat small intestine lysate

      Lysates/proteins at 10 µg per lane.

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

      Predicted band size: 48 kDa
      Observed band size: 46 kDa
      why is the actual band size different from the predicted?

    • Immunofluorescent staining of Cytokeratin 20 in SW480 cells, using ab109111 at a 1/50 dilution.

    References

    ab109111 has not yet been referenced specifically in any publications.

    Customer reviews and Q&As

    1-8 of 8 Abreviews or Q&A

    This product is known to not work in this application or species.
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citric acid
    Sample
    Rat Tissue sections (Bladder)
    Specification
    Bladder
    Permeabilization
    No
    Fixative
    Formaldehyde

    Mr. Carl Hobbs

    Verified customer

    Submitted Jan 13 2014

    This product is known to not work in this application or species.
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citric acid
    Sample
    Marmoset (common) Tissue sections (Small intestine)
    Specification
    Small intestine
    Permeabilization
    No
    Fixative
    Formaldehyde

    Mr. Carl Hobbs

    Verified customer

    Submitted Jan 13 2014

    This product is known to not work in this application or species.
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citric acid
    Sample
    Goat Tissue sections (Small intestine)
    Specification
    Small intestine
    Permeabilization
    No
    Fixative
    Formaldehyde

    Mr. Carl Hobbs

    Verified customer

    Submitted Jan 13 2014

    This product is known to not work in this application or species.
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citric acid
    Sample
    Human Tissue sections (Colon)
    Specification
    Colon
    Permeabilization
    No
    Fixative
    Formaldehyde

    Mr. Carl Hobbs

    Verified customer

    Submitted Jan 13 2014

    This product is known to not work in this application or species.
    Application
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
    Blocking step
    BSA as blocking agent for 10 minute(s) · Concentration: 1% · Temperature: 21°C
    Antigen retrieval step
    Heat mediated - Buffer/Enzyme Used: Citric acid
    Sample
    Dog Tissue sections (Small intestine)
    Specification
    Small intestine
    Permeabilization
    No
    Fixative
    Formaldehyde

    Mr. Carl Hobbs

    Verified customer

    Submitted Jan 13 2014

    Answer

    Thank you for your reply.
    I have processed the request to receive a vial of ab76126 as a free of charge replacement for ab10911. Your new order number is *.
    If there is anything else I can help you with, please let me know.

    Read More

    Answer

    Thank you for your reply.
    I am sorry that the protocol tips that you received back in February did not prove to be successful and I would be very happy to send you a replacement vial. However, instead of send another vial of ab109111, would you wish to try another antibody, against the same target? I have placed a link to antibody that may be suitable below:
    https://www.abcam.com/Cytokeratin-20-antibody-EPR1622Y-ab76126.html
    I look forward to your reply and helping resolve this issue.

    Read More

    Question

    DESCRIPTION OF THE PROBLEM Non-specific band SAMPLE SW480 and Colo320DM, colon cancer cell lines. PRIMARY ANTIBODY Ab I: Rabbit monoclonal Anti-Cytokeratin 20 (abcam 109111). Overnight incubation at +4 Celsius, dilution 1/10,000 in 3% non-fat dry milk in TBS-T. After incubation with the Ab I, I performed 3 washes of 5 minutes in TBS-T. DETECTION METHOD Pierce ECL Western Blotting substrate (32106) and Amersham ECL Prime western Blotting Detection reagents (RPN 2232). POSITIVE AND NEGATIVE CONTROLS USED SW480 positive control. ANTIBODY STORAGE CONDITIONS After arrival the Ab was stored at -20 Celsius. SAMPLE PREPARATION Whole proteins were extracted with lysis buffer [150 mM NaCl, 50 mM Tris/HCl, pH 8, 0.5 mM EDTA, 0.1 mM EGTA, 1% Triton X-100]with Roche's Complete Protease Inhibitor Cocktail Tablets. Samples were put in boiling water for 3 minutes. AMOUNT OF PROTEIN LOADED 50ug ELECTROPHORESIS/GEL CONDITIONS 8% SDS-PAGE TRANSFER AND BLOCKING CONDITIONS Whole proteins were extracted with cold lysis buffer (150 mM NaCl, 50 mM Tris/HCl, pH 8, 0.5 mM EDTA, 0.1 mM EGTA, 1% Triton X-100)incubating cells for 20 minutes on ice. Lysate were then centrifuged for 10 minutes at +4 Celsius at 7000xg and supernatant was transfer to a fresh tube and stored at -20 Celsius. SECONDARY ANTIBODY Ab II: Peroxidase conjugated goat anti-rabbit from Rockland (611-1322-0500). 45 minutes of incubation at room temperature, dilution 1/10,000 in TBS-T. After incubation with the Ab II, I performed 3 washes of 5 minutes in TBS-T. HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 3 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes WHAT STEPS HAVE YOU ALTERED? I tried to block in 3%BSA to eliminate aspecific and increased the concentration on Ab I because I am working with a CCD camera. I reduced the amount of non specif bands but i detected only a band around 120kDa!!!

    Read More
    Answer

    Thank you for contacting Abcam.
    I just have a few further questions for you, when you increased the primary antibody concentration from 1/10000, which was the primary antibody concentration that you used? I ask as it may be that you need to increase it further to see you band of interest. Also have to you tried to increase your boiling time from 3 minutes to 10 minutes, as the high molecular weight band that you are seeing could be your band of interest in a complex with other proteins. By boiling longer you may be able to break that complex up and so resolve you band at 48kDa.
    I look forward to your reply and helping you resolve this issue.

    Read More

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