Hi,
I am currently working on the Antibody phage library. It went well till phage development, but I am facing issues with the biopanning step. I am not getting binders even after 4rounds of panning. If I get colonies after binding, it is contaminated or no DNA observed.
Is there any way we can check the phages for production of ScFv before the biopanning round to assure?
Harshana,
Are your bacteria for producing free ScFv from the phage-DNA ok?
Otherwise, when you have amplified the library in the phage stage, before going into another round of screening, you could analyze an aliquot of your phages by SDS-PAGE. You should see the phage proteins on the gel and the modified coat protein should have the respective size.
Have you sequenced single clones of your library to assure the library plasmids have the expected sequence and elements?
Which system are you using?