Overview

  • Product name
    Anti-CD81 antibody [M38]
    See all CD81 primary antibodies
  • Description
    Mouse monoclonal [M38] to CD81
  • Host species
    Mouse
  • Tested applications
    Suitable for: WB, IP, IHC-P, Functional Studies, ICC, Flow Cytmore details
  • Species reactivity
    Reacts with: Rabbit, Cat, Human
  • Immunogen

    Tissue, cells or virus corresponding to Human CD81. MOLT4 cell line

  • Positive control
    • Human MOLT4 T cell line. This antibody gave a positive result in IHC in the following FFPE tissue: Human normal liver. For Western Blot: Jurkat cell line Flow Cyt: HAP1-WT cells.

Properties

Applications

Our Abpromise guarantee covers the use of ab79559 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 Use at an assay dependent concentration. Use under non reducing condition. Predicted molecular weight: 26 kDa.
IP Use at an assay dependent concentration.
IHC-P Use at an assay dependent concentration.
Functional Studies Use at an assay dependent concentration. In human MOLT4 T cell line ab79559 inhibits syncytium formation induced by coculture with human T-cell leukemia virus type 1 (HTLV 1)-positive human T-cell lines.
ICC Use at an assay dependent concentration.
Flow Cyt Use a concentration of 1 µg/ml.

ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.

 

Target

  • Function
    May play an important role in the regulation of lymphoma cell growth. Interacts with a 16-kDa Leu-13 protein to form a complex possibly involved in signal transduction. May acts a the viral receptor for HCV.
  • Tissue specificity
    Hematolymphoid, neuroectodermal and mesenchymal tumor cell lines.
  • Involvement in disease
    Defects in CD81 are the cause of immunodeficiency common variable type 6 (CVID6) [MIM:613496]; also called antibody deficiency due to CD81 defect. CVID6 is a primary immunodeficiency characterized by antibody deficiency, hypogammaglobulinemia, recurrent bacterial infections and an inability to mount an antibody response to antigen. The defect results from a failure of B-cell differentiation and impaired secretion of immunoglobulins; the numbers of circulating B cells is usually in the normal range, but can be low.
  • Sequence similarities
    Belongs to the tetraspanin (TM4SF) family.
  • Post-translational
    modifications
    Not glycosylated.
  • Cellular localization
    Membrane.
  • Information by UniProt
  • Database links
  • Alternative names
    • 26 kDa cell surface protein TAPA 1 antibody
    • 26 kDa cell surface protein TAPA-1 antibody
    • 26 kDa cell surface protein TAPA1 antibody
    • CD 81 antibody
    • CD81 antibody
    • CD81 antigen (target of antiproliferative antibody 1) antibody
    • CD81 antigen antibody
    • CD81 molecule antibody
    • CD81_HUMAN antibody
    • CVID6 antibody
    • S5.7 antibody
    • TAPA 1 antibody
    • TAPA1 antibody
    • Target of the antiproliferative antibody 1 antibody
    • Tetraspanin 28 antibody
    • Tetraspanin-28 antibody
    • Tetraspanin28 antibody
    • Tspan 28 antibody
    • Tspan-28 antibody
    • Tspan28 antibody
    see all

Images

  • Overlay histogram showing HAP1 wildtype (green line) and HAP1-CD81 knockout cells (red line) stained with ab79559. The cells were fixed with 4% formaldehyde (10 min)  and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab79559, 1µg/ml) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) presorbed (ab150117) at 1/2000 dilution for 30 min at 22°C.

    A mouse IgG1 isotype control antibody  (ab170190) was used at the same concentration and conditions as the primary antibody (HAP1 wildtype - black line, HAP1-CD81  knockout - grey line). Unlabelled sample was also used as a control (this line is not shown for the purpose of simplicity).

    Acquisition of >5,000 events were collected using a 50 mW Blue laser (488nm) and 530/30 bandpass filter.

    This antibody can also be used in HAP1 cells fixed with  80% methanol (5 min), , permeabilized with 0.1% PBS-Triton X-100 for 15 min under the same conditions.

  • Overlay histogram showing MOLT4 cells stained with ab79559 (red line). The cells were fixed with 4% paraformaldehyde and incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab79559, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in MOLT4 cells fixed with methanol (5 min) used under the same conditions.

    Please note that Abcam do not have data for use of this antibody on non-fixed cells. We welcome any customer feedback.
  • IHC image of CD81 staining in Human normal liver formalin fixed paraffin embedded tissue section, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab79559, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

     

    For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.

  • Flow cytometry analysis using human peripheral blood cells with ab79559. 

References

This product has been referenced in:
  • Langevin SM  et al. Balancing yield, purity and practicality: a modified differential ultracentrifugation protocol for efficient isolation of small extracellular vesicles from human serum. RNA Biol 16:5-12 (2019). Read more (PubMed: 30604646) »
  • Martins SA  et al. CD81 Promotes a Migratory Phenotype in Neuronal-Like Cells. Microsc Microanal 25:229-235 (2019). Read more (PubMed: 30714554) »
See all 13 Publications for this product

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Application
Immunocytochemistry/ Immunofluorescence
Sample
Human Cell (SH-SY5Y)
Permeabilization
Yes - 0.2% Triton-X
Specification
SH-SY5Y
Blocking step
BSA as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 4°C
Fixative
Paraformaldehyde

Abcam user community

Verified customer

Submitted Nov 09 2015

Answer

Thank you for getting back to me and for confirming the order details and the batch number.
I would like to reassure you that our Abpromise applies to your complaint since you purchased this product within the guarantee period. This means that in the event that a product is not functioning in the applications/species cited on the product data sheet (and the problem has been reported within 6 months of purchase) we will happily offer a credit note/refund to the value of the product purchased.
Though, you have kindly provided some details, it would be much appreciated if I could get some more information which would help me identify the source of the problem.
Could you provide some further details of the protocol used and complete the following form (attached as a word document). It would be much appreciated if you could attach an image to the response.
I am particularly interested in the followings:
- sample preparation,
- lysis buffer,
- positive control,
- loading control
Thank you for your understanding and co-operation in this matter. I look forward to hearing from you and hope to solve this problem as soon as possible.

Read More

Question

Hi there,



A couple of months ago I purchased your rabbit anti-CD81 (anti-TAPA-1) antibody (ab109201) which is guaranteed to detect human CD81 by western blot. I have been unable to detect CD81 in blots and wonder if I am doing something incorrectly as another one of your antibodies (ab79559) also failed to wprk in western blot.



I am using 2 cell lines (Huh7.5 and 293T cells) and I have shown by flow cytometry that both contain surface CD81:

- they both strongly bind another one of your anti-CD81 antibodies (ab79559 - flow cytometry and WB guaranteed);

- they both bind HCV E2 (the receptor of which is CD81);

- cells pre-incubated withab79559 no longer bind HCV E2 (as CD81 is blocked by your antibody).



I wanted to confirm the presence of CD81 by WB so performed SDS-PAGE withwhole cell lysates (from Huh7.5, 293T, Raji, Jurkat and other cell lines) using standard protocols, transferred proteins onto nitrocellulose, blocked for 2 h with 10 % milk, and probed overnight withab79559 at 1:1000 dilution in 5 % milk in TBS-T, followed by anti-mouse-HRP. I even got no signal, even when reprobed at 1:100 dilution (incidentally, the problem is not with the secondary as that works with other antibodies).



I assumed thatab79559 must detect a 'correctly folded' epitope as it works with flow cytometry and that this must be altered by SDS-PAGE so I tried your WB-specific anti-CD81 antibody (ab109201). WB was performed as above, but using an anti-rabbit-HRP secondary, and again I got no signal.





I then transiently transfected 293T, 293T/17 and Huh7.5 cells with myc-DDK-tagged CD81 on the CMV promoter (Origene RC217508), and after 48 h expression performed SDS-PAGE and WB using whole cell lysates of transfected and non-transfected cells. I then probed one blot with your WB-guaranteed anti-CD81 antibody (ab109201) and another blot with an anti-flag antibody. The anti-flag antibody strongly detected CD81 in lanes from CD81-transfected cells, but nothing in non-transfected lanes. Strangely, your anti-CD81 antibody detected nothing - even the transfected lanes had no signal.



I had begun to think that CD81 perhaps did not run into the gel during SDS-PAGE (as membrane proteins can sometimes be temperamental), but I am perplexed that I can detect tagged CD81 with the anti-tag antibody, but not with your anti-CD81. Please can you tell me if there is something that I have failed to do correctly for the detection of CD81 by WB so that I can test the antibody again.



Kind regards,

Read More
Answer

Thank you for your enquiry regarding ab109201 and for taking the time to provide some useful details of the experiments. I am very sorry to hear that you are having problems with this antibody.
As our Abpromise indicates, in the event that a product is not functioning in the applications/species cited on the product data sheet (and the problem has been reported within 6 months of purchase) we will happily offer a credit note/refund to the value of the product purchased.
Could you please provide the batch and order numbers for ab109201?
I look forward to hearing from you and hope to solve this problem as soon as possible.

Read More

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