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ReviewID: xxx AbID: 84814, Anti-DHPR alpha 1 antibody [S57-46] Rating: Inconclusive Image: Anti-DHPR alpha 1 antibody [S57-46] for Immunohistochemistry (Frozen sections) (Human) Sample: Species: Human Type: Tissue sections Specification: atrium Application: Application: Fixative: Paraformaldehyde Permeabilization: Yes Blocking step: BSA as blocking agent for 1 hour at 25°C Other detail: Dilution: 1/50 Incubation time: 12 hours at 4°C Diluent: PBS+BSA+serum Secondary Antibody: Name: Non-Abcam Antibody was used: goat anti mouse IgG2b488 Conjugation: Alexa Fluor® 488 Dilution: 1/500 Additional Data: Additional Notes: only see non-specifc binding
Asked on Apr 27 2012
According to our records, ab84814 was proving difficult to use in IHC-Fr and we were in contact in order to help resolve the issue.
Looking at our correspondence, it appears that we are awaiting more details in order to help us better understand the difficulties experienced.
Based on the information in your review, I feel that altering the following step in your protocol may help improve your results:
1) Trying another fixative might help to solve the problem as the PFA fixation might crosslink antibody binding sites.
2) You could also try omitting the permeabilization step if your sections are thin enough (10 um or less)
3) Doing a no primary control could help to determine if the secondary antibody contributes to the non-specific staining.
4) What is you order or PO number?
If you decide to follow these suggestions, please let me know if they are helpful. Additionally, this antibody is covered by our Abpromise, so if the suggestions do not help and the antibody was purchased within the past 6 months, I would be happy to discuss a refund or replacement with you to resolve this issue.
If the requested information has already been sent, it appears that it did not reach our Scientific Support team and we apologize for this inconvenience. In this case we would like to ask for the information again so that we can reach a resolution.
If the issue has already been settled, please let us know so that we can be assured that the problem has been solved to your satisfaction and update our records.
We wish you the best of luck with your research and look forward to a reply.
Answered on Apr 27 2012