We are having problems with the Pharmacia phage-display kit. We
have inserted our single chain antibody gene into the phagemid vector
and we are making phage but we are unable to isolate positive clones
by ELISA. It seems we are not getting our antibody incorporated into
the phage particle. We have gone through two rounds of panning and still
have no positive results by ELISA. Does anyone know of any other phagemid
vectors that are available that may work better or what may be our problem.
I have also transformed a non-suppressor E.coli strain with the phagemid
vector and I am able to isolate soluble antibody protein because I inserted
an amber codon between my antibody gene and gene 3 of the phage. Therefore
we do not think it is a problem with our construct or antibody sequence
but we are unable to isolate whole phage with our antibody attached to g3.
A few months ago I heard Carl Barbas at Scripps had a phagemid vector.
Does anyone know if he is willing to give out the vector or how we might
contact him? Any suggestions would be greatly appreciated.
crumbley at uxa.cso.uiuc.edu