Anti-Hsp90 antibody [16F1] (ab13494)

I would like to use the above-mentioned antibody as primary antibody for FACS staining of human and mouse cells. Because of that I'm curious if the Ab has been tested for FACS (I saw that it worked in ELISA, so might we assume that will it work in FACS as well?)? If not, might it be possible to get a small aliquot for test?

ANSWER:

Thank you for contacting us.

To our knowledge, tha antibody ab13494 has not been tested in flow cytometry. It may work for that application, on fixed and permeabilized samples, but we do not guarantee this. We do not have small samples for testing.

Have you considered the mouse monoclonal ab13492 (clone AC88), which is also available conjugated to PE (ab65171) and FITC (ab65173)?

I would like to test this antibody in fall armyworm (SF9 cells), an untested species.

ANSWER:

I am very pleased to hear you would like to accept our offer and test ab13494 in fall armyworm (SF9 cells). This code will give you 1 free primary antibody before the expiration date. To redeem this offer, please submit an Abreview for fall armyworm and include this code in the “Additional Comments” section so we know the Abreview is for this promotion. 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.

I am looking to buy an Hsp90 and Aha1 (aka Ahsa1) antibody. I was wondering if antibodies ab91259, ab83036, ab13492 would work in zebrafish?

ANSWER:

Thank you for contacting us.

Unfortunately, none of the antibodies you mentioned (ab91259, ab83036, ab13492) have been tested in zebrafish so far.

However, we have another HSP90 antibody that was tested in zebrafish: ab13494

(http://www.abcam.com/Hsp90-antibody-16F1-ab13494.html) - it would be guaranteed for this species as well as the listed tested applications.

For Aha1, we do not have an antibody tested in this species. But if you please could send me the protein sequence for zebrafish, I'd be happy to check homology and to see whether any of the Aha1 antibodies can be used for this species.

If the homology is about 85% or higher, you can use our testing discount program.

For UNTESTED species and/or applications, we have established a testing discount program. Here is a brief description of how it works:

The testing discount program is for customers who like to use an antibody/protein on an untested species/application. You would purchase the antibody at full price, test it and submit an Abreview with your data (positive or negative). On your next order you will receive a discount for ONE antibody at the full price (100%) of the antibody you have tested. The terms and conditions applicable to this offer can be found here: www.abcam.com/collaborationdiscount.

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

BATCH NUMBER 213690 ORDER NUMBER -- NOT SPECIFIED --

DESCRIPTION OF THE PROBLEM No signal or weak signal

SAMPLE BHK cell lysate (baby Humster)

PRIMARY ANTIBODY primary Ab 1t 1:1000 ans 1:500 dilution 1Hr incubation. washed X3 in T-PBS

DETECTION METHOD Detection with ECL

POSITIVE AND NEGATIVE CONTROLS USED Positive was the lysate, detected then with other Rabbit anti HSP90 from [another company] got good results

ANTIBODY STORAGE CONDITIONS 4deg

SAMPLE PREPARATION Lysate was prepared using regular RIPA buffer pH7.4. pepstatin , leueptin 10mg/ml and PMSF 1mg/ml No sample heating

AMOUNT OF PROTEIN LOADED 100ug

ELECTROPHORESIS/GEL CONDITIONS Reducing on 7% SDS-Page

TRANSFER AND BLOCKING CONDITIONS wet transfer (100V for 100min) Then wet transfered to NitroCel. membrane. Membrane was blocked 1Hr 5% milk in T-PBS. S, Washed X3 and developed with ChemiLumisescnce GE.

SECONDARY ANTIBODY 1hr at RT with Secundary anti Rat HRP conj. (1:10000) [another company]

HOW MANY TIMES HAVE YOU TRIED THE APPLICATION? 2 HAVE YOU RUN A "NO PRIMARY" CONTROL? No DO YOU OBTAIN THE SAME RESULTS EVERY TIME? Yes

ANSWER:

Thank you for contacting us for technical support, I'm sorry to hear you are experiencing problems with ab13494.

I think the fact that another antibody against Hsp90 worked well rules out the possibility that your samples express low levels of the protein or that the extraction of the protein is not adequate. I strongly believe that you will get a good signal by incubating the antibody overnight at 4C and maybe by diluting it less. The recommendation is 2ug/ml (so 1:500) however depending on the tissue type you made need a bit more, so I would recommend to concentrate more (though I think that by incubating overnight you should have a good signal with 1:500). We have used an Ecl+ detection system and if an Ecl kit is used the antibody may need to be more concentrated as it is less sensitive. Please check that the secondary works well with other primary antibodies, if the other anti Hsp90 antibody is not a rat monoclonal. If you still experience a problem with those changes please do not hesitate to contact me and I will offer you a refund or replacement if the antibody was bought in the last 90 days,

Best of luck with your next set of experiments,

What method of antigen retrieval was used when testing this antibody in IHC-P?

ANSWER:

Thank you for your enquiry.

I have added references for the antibody as used in IHC to the datasheet. You may refer to these articles for further information. I have included a link to the datasheet below for your convenience.

I hope this information helps. Please do not hesitate to contact us if you need anything further.