Affinity-based screening of combinatorial libraries using automated, serial-column chromatography.
We have developed an automated serial chromatographic technique for screening a library of compounds based upon their relative affinity for a target molecule. A "target" column containing the immobilized target molecule is set in tandem with a reversed-phase column. A combinatorial peptide library is injected onto the target column. The target-bound peptides are eluted from the first column and transferred automatically to the reversed-phase column. The target-specific peptide peaks from the reversed-phase column are identified and sequenced. Using a monoclonal antibody (3E-7) against beta-endorphin as a target, we selected a single peptide with sequence YGGFL from approximately 5800 peptides present in a combinatorial library. We demonstrated the applicability of the technology towards selection of peptides with predetermined affinity for bacterial lipopolysaccharide (LPS, endotoxin). We expect that this technology will have broad applications for high throughout screening of chemical libraries or natural product extracts.
Evans
D M
DM
PerSeptive Biosystems, Framingham, MA 01701, USA.
Williams
K P
KP
McGuinness
B
B
Tarr
G
G
Regnier
F
F
Afeyan
N
N
Jindal
S
S
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D001709:Biotechnology; D002846:Chromatography, Affinity / Q000295:instrumentation / Q000379:methods*; D015195:Drug Design; D006801:Humans; D008070:Lipopolysaccharides / Q000276:immunology; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry / Q000302:isolation & purification; D019151:Peptide Library; D001615:beta-Endorphin / Q000737:chemistry / Q000276:immunology
D000911:Antibodies, Monoclonal; D008070:Lipopolysaccharides; D009842:Oligopeptides; D019151:Peptide Library; D001615:beta-Endorphin