Anti-Adenovirus 抗体 [B025 / AD51] (ab7428)

Thank you for your inquiry regarding ab7428 (Mouse Anti-Adenovirus).
Indeed, this antibody does require pretreatment with a protease enzyme. The successful use of ab7428 in IHC-paraffin has been reported in a customer publication which used this same clone, the citation for which is included below. Protease treatment was reported to be 15 minutes prior to incubation for 1 hour with 1:1000 diluted primary antibody.
http://www.ncbi.nlm.nih.gov/pubmed/1401177 Maddox A, Francis N, Moss J, Blanshard C, Gazzard B. J Clin Pathol. 1992 Aug;45(8):684-8. PMID:1401177
Regarding the Ventana autostainer, we have no record of this antibody being used with that apparatus.
Below I have included our recommended protocols for antigen retrieval in formalin-fixed paraffin embedded sections.
Proteolytic antigen retrieval using trypsin
Reagents
Calcium Chloride, (CaCl2), 0.1 g
Trypsin (Sigma Type II), 0.1 g
Distilled Water, 100 ml
Sodium Hydroxide, (NaOH), 0.1 M
Method
Dissolve CaCl2 in distilled water and adjust the pH to 7.8 with NaOH. Store at 37C.
Dissolve trypsin in the CaCl2 solution.
Place sections pronase solution at room temperature and incubate for pre-determined optimum time (approximately 10 minutes).
Place sections in trypsin solution at 37°C and incubate for a pre-determined optimum time (approximately 20-30 minutes).
Wash in TBS and proceed with staining.

Proteolytic antigen retrieval using pronase
Reagents
Calcium Chloride, (CaCl2), 0.1 g
Pronase 0.1g
Distilled Water, 100 ml
Sodium Hydroxide, (NaOH), 0.1 M
Method
Dissolve CaCl2 in distilled water.
Dissolve pronase in CaCl2 solution and pH to 7.8 with NaOH.
Wash in TBS and proceed with staining.

END PROTOCOL
I hope this information helps. If this does not improve your results, please let us know and we can look into replacing or refunding the product for you.

I am glad finally the product gave you expected results.

Many thanks for your cooperation in this case.

Thank you for confirming these details and for your cooperation. The details provided enable us to closely monitor the quality of our products.

I am sorry this product did not perform as stated on the datasheet and for the inconvenience this has caused. As requested, I have issued a free of charge replacement with the order number 1180968.

To check the status of the order please contact our Customer Service team and reference this number.

Please note that this free of charge replacement vial is also covered by our Abpromise guarantee. Should you still be experiencing difficulties, or if you have any further questions, please do not hesitate to let us know.

I wish you the best of luck with your research.

PS: The product ab3648 is currently out of stock and is expected on 16th October; we will send you the vial next week.

Thank you for your email.

I am sorry to hear that the antibody did not work. The proteinase kInase K was the recommended antigen retrieval for this product so I am sorry there is nothing more we can suggest.

I can certainly send you a vial of ab3648 if you are interested.

Thank you very much for your cooperation in this case.

Thank you for your enquiry.

I am sorry to confirm that we do not provide expiry dates. Antibodies should be stable for at least 1 year if stored correctly, as described on the individual datasheets.

I would like to reassure you that ab7428 Adenovirus antibody is covered by our 6 month guarantee for IHC-P, ELISA and IHC-Fr.

For more details regarding our guarantee, please see the following page from our website:

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

I hope this will be helpful. If you have any further questions, please do not hesitate to contact me.

Thank you very much for providing details.

I have few more question to understand the problem better:

- Is this ab fails to stain at all or showed high background?
- Have you tested the positivity of lung section by any other method?
- Could you provide an image?

I have following further suggestions for protocol optimizations.

Antigen Retrieval step: this is a very tricky step. Many antibodies needs optimizations so we always recommend our customers doing the troubleshooting for this step. In the following publication author have actually used protease/ enzyme based antigen retrieval system. I would suggest trying the similar AR procedure.http://www.ncbi.nlm.nih.gov/pmc/articles/PMC495144/pdf/jclinpath00422-0042.pdf.
Heat mediated antigen retrieval system can also be tried - Microwave with citrate buffer at pH 6.0. Please incubate the sections at a range of time e.g. 5 min, 10 min and 15 min. This range will help getting the optimized results.
Overnight incubation at 4C is recommended.
Please try normal serum or 5% BSA for 1-2 hour as blocking agent.
Wash the slides with PBST



I hope these suggestions will be helpful. Should the results do not improve please do not hesitate to contact me.

Thank you for contacting us.

I am sorry to hear you are experiencing difficulties with one of our products. We take product complaints very seriously, and investigate every product that we feel may not be performing correctly.

I am attaching our questionnaire so that we can gather further information regarding the samples tested and the protocol used. Once we receive the completed questionnaire, we will look at the protocol and see if there are any suggestions we can make that may improve the results. This information will also allow us to investigate this case internally and initiate additional testing where necessary. If the product was purchased in the last six months and is being used according to our Abpromise, we would be happy to replace or refund the antibody.

I look forward to receiving your reply.

Thank you for contacting us.

The information we have on the datasheet is from different publications; we unfortunately as such do not hold any tonsil related data. I apologize for being not helpful due to lack of data. I can recommend searching similar kind of staining in publications. I have attached few publications where authors have done Immunohistochemistry staining of Adenovirus in tonsil tissues. These publications might provide further insight in your research.

http://jcm.asm.org/content/48/2/642.full?maxtoshow=&hits=10&RESULTFORMAT=&fulltext=%25DF&searchid=1&FIRSTINDEX=963&resourcetype=HWFIG

http://jdc.jefferson.edu/cgi/viewcontent.cgi?article=1023&context=mifp&sei-redir=1&referer=http%3A%2F%2Fwww.google.co.uk%2Furl%3Fsa%3Dt%26rct%3Dj%26q%3Dadenovirus%2520staining%2520in%2520tonsil%26source%3Dweb%26cd%3D7%26sqi%3D2%26ved%3D0CF4QFjAG%26url%3Dhttp%253A%252F%252Fjdc.jefferson.edu%252Fcgi%252Fviewcontent.cgi%253Farticle%253D1023%2526context%253Dmifp%26ei%3DXOP6T_unBsrasga01-WSBQ%26usg%3DAFQjCNHCzteXZho2cf-gfNq54HymHXjlMQ#search=%22adenovirus%20staining%20tonsil%22

http://www.naro.affrc.go.jp/org/niah/disease/em/em_en/virus0.html

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

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Thank you for contacting us and reporting your results with using Anti-Adenovirus antibody [B025 / AD51] ab7428 in ELISA.
I have had a look into the data we have for this antibody. Unfortunately, no epitope mapping has been performed with this antibody and we can therefore not be sure of which part of the virus this antibody binds. We also have no data on if the antibody can recognise other serotypes to the one which it was raised (3). I am sorry that I could not provide any better information.
If you would like me to look at your ELISA protocol, as discussed over the phone, to see if I can suggest any improvements to increase the signal observed, please could you fill out the questionnaire I have attached to this email.
I am currently having a look into which antibodies we have which can be said to recognise both serotype 3 and 11 for your ELISA. Currently the information I have is that ab35546 (unconjugated) and ab20021 (FITC conjugated) are both able to recognise serotypes 3 and 11 and both have been used in ELISA previously.
I am currently looking into a few other antibodies which may also be able to recognise both forms and will get back to you when I have more information.
I hope this information has been of help. If you have any further questions please do not hesitate to contact us again.

Thank you for your enquiry. Unfortunately I do not have details of the degree of reactivity of ab8249 and ab8251 against the Adenovirus subclasses. However, I can tell you that ab7428 recognises all Adenovirus serotypes. All antibodies are yet to be tested by neutralisation. All tested applications are specified on Abcam product datasheets. At Abcam we are always interested in determining new applications of our antibodies. Therefore I would like to offer you a discount of 20% off the price of ab8249 in exchange for a review of this antiserum by neutralisation. Please e-mail me if you are interested and I will supply you with a unique discount code that you can enter during the ordering process. I look forward to hearing from you.