I'm developing an ELISA for coccidian parasite Cystoisoispora. I'm wondering if I precipitate proteins of sporozoite/merozoite lysate by TCA I can be sure of quality or I'd better use other concentration methods?
I wouldn't use protein precipitation methods. After TCA you will loose native structure of your antigen. For Western Blot is does not matter but here it will be problem.
You can use the ammonium sulfate precipitation and then a dialysis. it is an old method but it works well and it does not denature your proteins. It depends on the amount of proteins you have and the starting volume....
I agree. Ammonium sulphate precipitation and dialysis might work. But if you have many samples it will be problematic. On one well (96 well plate) you can load up to 250 ul of your sample. Is it still not enough to have acceptable signal? Maybe you should modify your ELISA protocol.
Hi I have prepared antigens using this system with much success in Sandwich elisa. Hope this helps
@Marie-José thanks for recommendation. @Dawid I would prefer to use much more zoites (although though to get enough material) for higher protein, however regarding loading 250 ul of antigen to each well do you think it is acceptable for reviewers, since I've not seen this high amount in any article. This question is just for my info!
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