I’ve been optimizing an ELISA setup, but the antibodies I’ve been using either show weak binding or high background. It’s driving me crazy because I’ve wasted so much sample already. Can anyone recommend how to find antibodies that actually perform well in ELISA without endless trial and error?
Totally feel your pain — optimizing ELISA is a delicate process, and a weak antibody can ruin everything. I had similar issues where the binding was poor, and the background was all over the place. A postdoc in my lab suggested checking out Gentaur https://gentaur.co.uk/antibodies/ , and that was a turning point. They actually label which antibodies have been validated for ELISA, so you’re not guessing anymore. The pair I got from them for detecting cytokines worked beautifully — clean signal, minimal noise, and consistent results between runs. I think their attention to validation and purity levels makes a huge difference. Now I use their catalog as my first stop before testing anything new.
I can totally relate to your ELISA story! The first time I ran an ELISA test, my readings were all over the place. Later I found out my washing buffer was too harsh and was stripping off the capture antibody. Now I double-check all my reagents before blaming the antibody. I think once you get the protocol locked in, even mid-tier antibodies can perform well if handled right. Still, nothing beats having a solid validated product — saves so many headaches.