Anti-HLA Class I antibody [W6/32] - Low endotoxin, Azide free (ab23755)

Overview

  • Product name
    Anti-HLA Class I antibody [W6/32] - Low endotoxin, Azide free
    See all HLA Class I primary antibodies
  • Description
    Mouse monoclonal [W6/32] to HLA Class I - Low endotoxin, Azide free
  • Host species
    Mouse
  • Specificity
    The antibody recognises an antigenic determinant common to HLA A, B and C. In WB, it detects the heavy chain of MHC molecule. We have data to indicate that this antibody may not cross react with Rabbit. However, this has not been conclusively tested and expression levels may vary in certain cell lines/tissues.
  • Tested applications
    Suitable for: Flow Cyt, IP, WB, IHC-Fr, ELISA, ICC/IF, Functional Studiesmore details
  • Species reactivity
    Reacts with: Cow, Cat, Human
  • Immunogen

    Tissue/ cell preparation - Membrane of human tonsil cells.

  • General notes

    The antibody recognises virtually all nucleated human cells, it is a valuable reagent for analysing variations in HLA class I expression in different disease states e.g. liver disease, muscular dystrophy, inflammatory myopathy and other neuromuscular disorders. This antibody is also suitable as a positive control for HLA tissue typing and crossmatching.

Properties

  • Form
    Liquid
  • Storage instructions
    Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term.
  • Storage buffer
    Preservative: None
    Constituents: PBS, pH 7.4
  • Concentration information loading...
  • Purity
    Immunogen affinity purified
  • Primary antibody notes
    The antibody recognises virtually all nucleated human cells, it is a valuable reagent for analysing variations in HLA class I expression in different disease states e.g. liver disease, muscular dystrophy, inflammatory myopathy and other neuromuscular disorders. This antibody is also suitable as a positive control for HLA tissue typing and crossmatching.
  • Clonality
    Monoclonal
  • Clone number
    W6/32
  • Isotype
    IgG2a
  • Research areas

Applications

Our Abpromise guarantee covers the use of ab23755 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 1µg for 106 cells.

Paraformaldehyde or methanol fixed cells.

ab170191 - Mouse monoclonal IgG2a, is suitable for use as an isotype control with this antibody.

IP Use at an assay dependent concentration.
WB 1/250. Use under non reducing condition. Detects a band of approximately 45 kDa (predicted molecular weight: 31 kDa).
IHC-Fr Use at an assay dependent concentration.
ELISA Use at an assay dependent concentration.
ICC/IF Use a concentration of 1 µg/ml.
Functional Studies Use at an assay dependent concentration.

Target

Images

  • Overlay histogram showing Jurkat cells stained with ab23755 (red line). The cells were fixed with 4% paraformaldehyde (10 min) 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 (ab23755, 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 IgG2a [ICIGG2A] (ab91361, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Jurkat 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.
  • ICC/IF image of ab23755 stained HeLa 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 (ab23755, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 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.

References

This product has been referenced in:
  • Wu H  et al. IFN-t Mediated Control of Bovine Major Histocompatibility Complex Class I Expression and Functionviathe Regulation of bta-miR-148b/152 in Bovine Endometrial Epithelial Cells. Front Immunol 9:167 (2018). Read more (PubMed: 29456541) »
  • Tong M  et al. Quantitative Analysis of Cellular Proteome Alterations in CDV-Infected Mink Lung Epithelial Cells. Front Microbiol 8:2564 (2017). Read more (PubMed: 29312244) »
See all 11 Publications for this product

Customer reviews and Q&As

1-10 of 19 Abreviews or Q&A

Application
Immunoprecipitation
Sample
Human Cell lysate - whole cell (TE-11 cells)
Total protein in input
10 µg
Immuno-precipitation step
Protein A
Specification
TE-11 cells

Abcam user community

Verified customer

Submitted Oct 27 2017

Application
Flow Cytometry
Sample
Human Cell (TE11, PBMC)
Permeabilization
No
Gating Strategy
HLA Class I positive living cells
Specification
TE11, PBMC
Fixation
None

Abcam user community

Verified customer

Submitted Nov 09 2015

Answer

I have received a reply from the lab who confirm that we have tested only cell surface binding of this antibody (which was good), not intracellular binding.

Currently we do not have data regarding its reactivity with premature antigen. However, for your experiments I suggest to use also other similar transmembrane proteins as a positive control to check the GA compartment is accessible well for the antibody staining.

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Answer

Thank you for contacting us.

To our knowledge our HLA Class I antibodies have not been tested to not detect HLA-G. I did find that our anti-HLA Class I [MEM-123] antibodies have the following specificity:

This antibody reacts with all human classical MHC Class I molecules in native cell-surface forms as well as with human HLA-G cDNA transfected cells. This antibody completely blocks the binding of pan MHC antibody (ab7855) but does not block binding of anti-HLA-G (ab7758/7904) to surface-expressed HLA-G.

While many of the HLA Class I products are created from tissue preparation and have not been epitope mapped, I have contacted the lab to determine whether the peptide immunogen used to create ab134743 is from a region which differs from HLA-G. I will let you know the result of that query.

Because we carry over 70,000 products, it isn't feasible for us to keep small sample sizes of our products. We are happy to reassure our customers that all of our products are covered by our Abpromise, which guarantees that the product will work in the applications and species specified on the datasheet, or we will offer a replacement, credit, or refund within 6 months of purchase.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

More information about our MEM-123 clone antibodies can be found at the following links:
https://www.abcam.com/hla-class-i-antibody-mem-123-ab2217.html
https://www.abcam.com/HLA-Class-I-antibody-MEM-123-Phycoerythrin-ab58998.html

Our ab134743 datasheet is at the following location:
https://www.abcam.com/HLA-Class-I-antibody-ab134743.html


Use our products? Submit an Abreview. Earn rewards!

https://www.abcam.com/abreviews

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Answer

Thank you for your enquiry.

I am sorry to confirm that this antibody is not tested and covered by our 6 monthy guarantee for this particular application. For more details regarding our guarantee, please see the following page from our website:

https://www.abcam.com/index.html?pageconfig=abpromise

Regrettably, as this is not currently a standard application, we are not able to offer our testing discount on this occasion.

However, I am pleased to provide a 5% discount off your purchase of this antibody (ab23755), which I hope will be helpful. In order to obtain this discount, please use the following code when you place the order:

5% Discount code: 5PC-OFF-T7T62

Also, you can obtain further reward by submitting an Abreview about this product via the online product datasheet. We always encourage customers to send their results back to us, whether positive or negative, and we make all product information available to researchers.

To find out more about our Abreview system and the Abpoints rewards you can obtain, please see the following webpage:

https://www.abcam.com/abreviews

I hope this will be helpful. Thank you for your time and effort.

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Answer

Thank you for contacting us.





The reactivity with bovine material is good. With regard to this product's cross reactivity with murine material please see the article referenced below:



Kievits F, Ivanyi P: Monomorphic anti-HLA monoclonal antibody (W6/32) recognizes polymorphic H-2 heavy-chain determinants exposed by association with bovine or human but not murine beta 2-microglobulin.Hum Immunol. 1987 Oct;20(2):115-26.





I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

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Answer

Thank you for contacting us.

Yes, we did offer that customer a discount to test the antibody in rat in IHC-P. We have a testing discount program where customers can purchase an antibody risk-free to try in their novel species and/or application. It's essentially buy-one-get-one free where you purchase the antibody, test it in the untested species or application, submit an Abreview with the data whether positive or negative, and get a discount code for 1 free antibody off a future purchase within 4 months.

Unfortunately it appears that this customer never got back to us with the data, so I do not know his/her results.

I hope this information is helpful to you. Please do not hesitate to contact us if you need any more advice or information.

Use our products? Submit an Abreview. Earn rewards!
https://www.abcam.com/abreviews

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Question
Answer

DISCOUNT CODE: *********
Expiration date: *********

I am very pleased to hear you would like to accept our offer and test ab23755 in mouse tissue. This code will give you: 1 free PRIMARY ANTIBODY before the expiration date. To redeem this offer, please submit an Abreview for mouse tissue and include this code in the “Additional Comments” section so we know the Abreview is for this promotion. For more information on how to submit an Abreview, please visit the site: www.abcam.com/Abreviews.


Remember, we publish both positive and negative Abreviews on our datasheets so please submit the results of your tests. The code will be active once the Abreview has been submitted and can be redeemed in one of the following ways: 1) Call to place your order and mention the code to our customer service department; 2) Include the code in your fax order; 3) Place your order on the web and enter the promotional code.

Any feedback that you can provide will be greatly appreciated, whether positive or negative. If you have any further questions, please do not hesitate to contact us. We look forward to receiving your Abreview and wish you luck with your research.

The terms and conditions applicable to this offer can be found here: www.abcam.com/collaborationdiscount.

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Answer

Thank you for your response. Lysates from Jurkat or HeLa cells or from human tonsil tissue can be used as positive control. Currently, we have do not have HLA Class I over-expression cell lysate in our catalogue. It may be worth optimizing the working concentration/dilution of this antibody to get stronger band on the blot. I hope this helps and if I can assist further, please do not hesitate to contact me.

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Answer

Thank you for your enquiry and your interest in our products. Anti-alpha tubulin (clone TU-01) can be used as loading control together with antibody HLA Class I (ab23755) . Currently, we have several forms of this antibody ie. unconjugated as well as conjugated: - ab7750 - Anti-alpha Tubulin antibody [TU-01] - ab18247 - Anti-alpha Tubulin antibody [TU-01] (FITC) - ab46708 - Anti-alpha Tubulin antibody [TU-01] (Dyomics® 547) - ab74696 - Anti-alpha Tubulin antibody [TU-01] (Biotin) - ab78108 - Anti-alpha Tubulin antibody [TU-01] I hope this helps and if I can assist further, please do not hesitate to contact me.

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1-10 of 19 Abreviews or Q&A

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