Anti-VIP 抗体 (ab8556)

The stock solution of the Optiblot DTT Reducer (10X) ab119199 in 2M. This means that once diluted 10 times this produces a 200 mM solution. Therefore if you usually use 10 mM with your samples I would suggest diluting this 200 times. The higher concentration should not really have an effect of the running of protein gels but as the pKa is around 8, in high concentration it can alter the pH somewhat which may affect the migration. We would normally suggest using 50 mM to ensure fully reducing the disulphide bonds in your samples.

Thanks for keeping me updated.

I'm sorry to hear that the flow cytometry results were not so good. We haven't tested this antibody in flow so I don't have specific recommendations for a protocol, and it might requrie some optimization as you've said. Do you know where in the cell the VIP accumulates before it is secreted? Have you considered fixing after staining with the primary antibody, or not fixing at all? Fixation might not be necessary if you're analyzing the cells immediately after staining.

I look forward to hearing from you about this and the IHC results with acetone-fixed tissue. Have a great day!

Thanks for your reply. After further discussion with the lab, I don't think I'll find out anything else about the IHC-Fr protocol, so I would recommend going ahead and trying the acetone fixation and some of the other alterations. If you do use up the antibody while troubleshooting, I'll be happy to send another vial, so don't worry about that.

How did the flow cytometry turn out?

Please let me know if you have any questions. Have a great day!

Thanks again for your patience.

I've heard back from the lab, and they did not test ab8556 in-house on frozen tissue sections. This application was added to our datasheet based on feedback from an outside lab, and I haven't tracked down information about the IHC-Fr protocol used yet. I will remove IHC-Fr from our datasheet if I can't find more information, and I apologize for this inconvenience. My contact at the lab did recommend a fixation with ice cold acetone for 10 minutes, so you could try this fixation if you would like.

Have you had a chance to try any of the methods to reduce the auto-fluorescence in the tissue, or have you gotten any other results with positive control tissue? This application is still covered by our Abpromise guarantee, so I will be happy to send a replacement product or issue a credit or refund if we can't get the antibody to work as it should.

I look forward to hearing from you. Please let me know if you have any further questions or if there is anything else that I can do for you.

Thank you for your call yesterday and for letting us know about the trouble with ab8556 in immunohistochemistry.

I apologize for the delay getting back to you,as I'm still waiting for more information from the lab regarding the suggested fixative and dilution for IHC on frozen tissue sections. As we discussed, antigen retrieval can be performed on frozen tissues, and we recommend the article by Yamashita, et al for a good method-

http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=Search&Term=%22Yamashita%20S%22%5BAuthor%5D&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_RVAbstractPlusDrugs1, http://www.ncbi.nlm.nih.gov/sites/entrez?Db=pubmed&Cmd=Search&Term=%22Okada%20Y%22%5BAuthor%5D&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_RVAbstractPlusDrugs1. Application of heat induced antigen retrieval to aldehyde fixed fresh frozen sections. http://www.jhc.org/. 2005 Nov;53(11):1421-32.

Theyautoclaved tissue sections in pH 9 Tris Hcl for 10 minutes at 120C. Ithink the increased pressure helps the tissue adhere to the slides.

I've also attached a great guide that I found regarding autofluorescence, the various sources, and ways to fix the problem. Theauthorsdon't really discuss glycine treatment but there are several other recommendations that would be helpful.

I will get back to you as soon as I have more information from the lab. I am sorry for the delay, but please let me know if you have any questions in the meantime or if there is anything else that I can do for you.

DISCOUNT CODE: xxxx Expiration date: xxxx I am very pleased to hear you would like to accept our offer and test ab8556 in Flow Cytometry. This code will give you 1 free PRIMARY ANTIBODY before the expiration date. To redeem this offer, please submit an Abreview for Flow Cytometry 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.

Thank you very much for your interest in ab8556. To our knowledge, ab8556 has not been tested in Flow Cytometry. Therefore, I can offer a discount off a future purchase if you buy ab8556 now, test it in Flow Cytometry and submit feedback to us in the form of an Abreview. It doesn’t matter whether the Abreview is positive or negative, we would just really like to receive your feedback. The discount would be to the value of 1 free PRIMARY ANTIBODY. If you are interested in this offer, please follow these steps: 1. Reply to this e-mail to let me know that you would like to proceed and test ab8556 in Flow Cytometry. I will then send a discount code. This code must be issued before purchasing ab8556 so please wait for my reply before ordering. 2. Purchase ab8556 either by phone, fax, or online (www.abcam.com). 3. Test it in Flow Cytometry. 4. Let us know the results, positive or negative, using our Abreview system (this will take about 10 minutes and images are great if you have them!). To find out how to submit an Abreview, please visit: https://www.abcam.com/abreviews. 5. After the review is submitted to us, the discount code becomes active. Simply place your new order by phone, fax, or on the web and mention the discount code. The discount can be redeemed for any PRIMARY ANTIBODY ordered and the discount code is valid for 4 months after issue. We are always pleased to obtain feedback about our products and any information is greatly appreciated! Even if ab8556 turns out to be unsuitable for Flow Cytometry, you will still receive the discount on your next purchase after your Abreview has been submitted. Please let me know if you have any questions about this offer and I would be happy to help you further. The Terms and Conditions of this offer can be found at: www.abcam.com/collaborationdiscount.  

Thank you for contacting me regarding this. It is important to have the code prior to ordering the first vial and to include that code as a reference in either your fax or email order, or to tell the customer service representative the code. They will use that to reference the Abreview when you place the subsequent order. Please let me know if you have any questions.