Hello, I want to detect by ELISA the presence of an administrated anti-Abeta antibody in mice brains. The mice were treated with a recombinant humanized antibody intraperitoneally and I want to know if (and how much) antibody would have crossed the BBB.
My doubts concern about the most appropriate lysis protocol of the brains for detecting those antibodies by ELISA.
If there is a more adequate way to perform the experience, I am open to advises and comments.
Thank you very much!
NP-40 lysis protocol for mouse brain. A2B5 is a neoepitope antibody for Abeta staining.
Thank you for your Answer, Mr. Boyao, but I am not trying to detect Abeta. I´m trying to determine the quantity of anti-Abeta antibody I injected intraperitoneally actually crossed the BBB and entered the brain. It is not a "normal IgG", it is modified with another functional group that could make easier BBB crossing and I want to know if it is working.
You can homogenate the brains in PBS. This will not interfere wiht the assay. If you use any SDS or other detergents in the lysis buffer which is probably the better option to get better recovery, you will have to make a serial dillution and test the amount of detergent that no longer intereferes with your ELISA assay. Hope ths will help!
Hello Steffen, thank you for your answer. Yes, I would use PBS and maybe a low percentage of TX100... I am thinking about using a protein G column to isolate the antibody and then concentrate the elution to increase the concentration before the detection by ELISA.
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