Anti-HLA E antibody [MEM-E/08] (ab11821)

In the data sheet for the HLA-E (MEM-E/08) antibody following crossreactivity is mentioned: The published results revealed that antibody is remarkably specific for HLA E, only with weak cross-reactivity with following classical MHC Class I molecules: HLA A24, B7, B27, B51, B54, C7 Unfortunatly there are no literature references provides to get informed how much cross reactivity is effecting results. Could you send me the literature references?

ANSWER:

Thank you for your patience in this matter.

The originator of ab11821 (HLA E antibody [MEM-E/08]) has provided the following references:

Sibilio L. et al., Tissue Antigens 62, 339, 2003. (Abstract 45 from the 3rd Int. Conference on HLA-G, Paris, July 2003).

Palmisano GL, Contardi E, Morabito A, Gargaglione V, Ferrara GB, Pist illo MP.: HLA-E surface expression is independent of the availability of HLA class I signal sequence-derived peptides in human tumor cell lines. Hum Immunol. 2005 Jan;66(1):1-12.

I hope this helps.

I am trying to titrate the antibody but need the appropriate cells/conditions, because HLA-E is not constitutively expressed on the surface of all cells. I tried fibroblasts and saw some slight internal staining, but only at the 1:12.5 dilution, which is very weak. Once I establish the correct titer to use, I can test the antibody for cross reactivity with rhesus macaque cells, the ultimate goal.

How was this antibody tested for application in FACS? What dilution and samples were used?

ANSWER:

This antibody was tested on the human cell line LCL 721.221 (HLA-E transfected), dilution 1:50 - 1:500. This antibody reacts with the native molecule of HLA-E.

I hope this helps. Please contact us again if you have any additional questions.

It is believed that HLA-E is universal expressed on lymphocyte (Human Immunology 1990 (29),131 & Tissue Antigen 1999 (53),292). I used primary aortic endothelial cells and peripheral lymphocytes to test this antibody. Both of them showed negative results. I don't know it is because the expression level is too low to see or other reasons. Did you test this antibody on lymphocyte?

ANSWER:

Thank you for your enquiry.

This antibody perfectly works in flow cytometry but as far as we know, this antibody was not tested on lymphocytes. It was tested in FACS with HLA-E transfected human cell line LCL 721.221.

Have you tried titrating the antibody? We generally recommend a starting point of 1ug/ml per 1x10^6 cells. However, since we don't have a recommended dilution on the datasheet, it would be important to optimize.

I hope this information helps, please do not hesitate to contact us if you need any more advice or information.

What types of cells has this antibody been tested on in FACS?

ANSWER:

Thank you for your equiry.

For testing this antibody on FACS was used human cell line LCL 721.221, HLA-E transfected.

Please do not hesitate to contact us if you need anything further.

I wish to use the above antibody for Flow cytometry on HeLa lines, but am unsure at what levels HLA-E is expressed on these cells. Do you have any data on the level of HLA-E expression demonstrated by HeLa cells?

ANSWER:

Thank you for your enquiry and your interest in our products.

This antibody perfectly works in flow cytometry but we have not used HeLa cell line for this study. Perhaps this information can be found in some published paper.