Trouble in phage display - (Jul/17/2012 )

hi,

i am having quite a serious problem with my phage display. i am working on human phage display tech and after panning as per the protcol, i cant express my scfv

clones. it is quite frustating as i am stuck with this for pretty long time...i will be very thankful for any help In detail about my experiment

1). Immobolise the Antigen (dissolved in coating buffer)

2). perform panning (incubate the phage library for one hour and collect the unbound phage and wash the wells extensively with TBS-T buffer and then add log phase e.coli and incubate for 20 mins and collect the infected e.coli and spread on LB-A plate

3). collect the well isolated colonies and perform PCR to pick the positive clone and then express the PCR positive clone with glucose suppression.

i am unable to express the PCR positive clones.....

i will be very thankful for any suggestion

You can try in vivo phage display, it can contain peptide native sturcture.