Towards proteome scale antibody selections using phage display.
In vitro antibody generation by panning a large universal gene library with phage display was employed to generate antibodies to more than 60 different antigens. Of particular interest was a comparison of pannings on 20 different SH2 domains provided by the Structural Genomics Consortium (SGC). Streamlined methods for high throughput antibody generation developed within the 'Antibody Factory' of the German National Genome Research Network (NGFN) were demonstrated to minimise effort and provide a reliable and robust source for antibodies. For the SH2 domains, in two successive series of selections, 2668 clones were analysed, resulting in 347 primary hits in ELISA. Half of these hits were further analysed, and more than 90 different scFv antibodies to all antigens were identified. The validation of selected antibodies by cross-reactivity ELISA, western blot and on protein microarrays demonstrated the versatility of the in vitro antibody selection pipeline to generate a renewable resource of highly specific monoclonal binders in proteome scale numbers with substantially reduced effort and time.
Mersmann
Michael
M
Technische Universität Braunschweig, Institute of Biochemistry and Biotechnology, Braunschweig, Germany.
Meier
Doris
D
Mersmann
Jana
J
Helmsing
Saskia
S
Nilsson
Peter
P
Gräslund
Susanne
S
Colwill
Karen
K
Hust
Michael
M
Dübel
Stefan
S
D000906:Antibodies / Q000276:immunology* / Q000302:isolation & purification*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D019151:Peptide Library; D020543:Proteome / Q000276:immunology* / Q000302:isolation & purification*
D000906:Antibodies; D019151:Peptide Library; D020543:Proteome