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Thank you for your email. I think the main problem is that the antibody is raised using an E. coli cell extract. The antibody works in the described ELISA assay procedure with a cell suspension dried on the plate. Accordingly, there is no guarantee that the antibody would recognize free E. coli cell suspension. I am also concerned how fully antigen recognition is maintained upon biotinylation.
I was wondering if you could provide me with free evaluation samples of the antibody you have against E. coli before biotinylation and I will try to use protein A-coated beads to capture the antibodies (instead of the streptavidin-coated beads I have been using). Since the polyclonal antibody is purified using protein A, that should improve the chances. I am using a modified form of E. coli K-12 strain for the work I am doing. The idea is to try to capture the cells in a suspension on paramagnetic beads to which E. coli antibody has been previously attached.
I hope you will be able to provide me with samples of your antibodies recommended by your colleagues. Hopefully, one of them will work in my system. Then, I will purchase the right antibody for continuation of our studies.
Your cooperation will be greatly appreciated.
Asked on Jul 06 2012
Thank you very much for your reply.
At this time, we do not have the unconjugated format of ab68451 in our catalog, but if you are interested in trying it Iwill see if we can add this antibodyto ourcatalog. We also have two unconjugated goat polyclonal antibodies, ab13627 and ab25823, which may be suitable as well-
Protein G beads should be used with the goat antibodies, as goat IgG has little affinity to protein A.
We only have full size vials of all of our antibodies, so unfortunately we do not have free samples to send out. These antibodies have not been tested ina bacterial pulldown assay, but if you would like to try one of these other unconjugated E. coli antibodies, we do have a testing discount program that you may be interested in. As part of this program, you would initially purchase the unconjugated E. coli antibody, test it in your assay, then submit an Abreview with your data and protocol. We will then give you one free primary antibody on a future purchase. I can send more information about this program if you would like.
Please let me know if you are interested in any of these other E.coli antibodies or if you have further questions, and I'll be happy to help.
Answered on Jul 06 2012