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J Lab Autom. 2012 Jun;17(3):186-200. doi: 10.1177/2211068211433503. Epub 2012 Feb 14.

Process automation toward ultra-high-throughput screening of combinatorial one-bead-one-compound (OBOC) peptide libraries.

Cha J, Lim J, Zheng Y, Tan S, Ang YL, Oon J, Ang MW, Ling J, Bode M, Lee SS.

Source

Institute of Bioengineering and Nanotechnology, Singapore.

Abstract

With an aim to develop peptide-based protein capture agents that can replace antibodies for in vitro diagnosis, an ultra-high-throughput screening strategy has been investigated by automating labor-intensive, time-consuming processes that are the construction of peptide libraries, sorting of positive beads, and peptide sequencing through analysis of tandem mass spectrometry data. Although instruments for automation, such as peptide synthesizers and automatic bead sorters, have been used in some groups, the overall process has not been well optimized to minimize time, cost, and efforts, as well as to maximize product quality and performance. Herein we suggest and explore several solutions to the existing problems with the automation of the key processes. The overall process optimization has been done successfully in orchestration with the technologies such as rapid cleavage of peptides from beads and semiautomatic peptide sequencing that we have developed previously. This optimization allowed one-round screening, from peptide library construction to peptide sequencing, to be completed within 4 to 5 days. We also successfully identified a 6-mer ligand for carcinoembryonic antigen-cell adhesion molecule 5 (CEACAM 5) through three-round screenings, including one-round screening of a focused library.

PMID:

22357565

[PubMed - indexed for MEDLINE]

Publication Types, MeSH Terms, Substances

Publication Types

Research Support, Non-U.S. Gov't

MeSH Terms

Automation, Laboratory*

Carcinoembryonic Antigen/chemistry

Combinatorial Chemistry Techniques/instrumentation

Combinatorial Chemistry Techniques/methods

Diagnosis

GPI-Linked Proteins/agonists

GPI-Linked Proteins/chemistry

High-Throughput Screening Assays*

Humans

Ligands

Microspheres

Peptide Fragments/chemistry

Peptide Fragments/isolation & purification*

Peptide Library*

Sequence Analysis, Protein/methods

Tandem Mass Spectrometry

Tumor Markers, Biological/agonists

Tumor Markers, Biological/chemistry

Substances

CEACAM5 protein, human

Carcinoembryonic Antigen

GPI-Linked Proteins

Ligands

Peptide Fragments

Peptide Library

Tumor Markers, Biological

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Process automation toward ultra-high-throughput screening of combinatorial one-b...
Process automation toward ultra-high-throughput screening of combinatorial one-bead-one-compound (OBOC) peptide libraries.
J Lab Autom. 2012 Jun ;17(3):186-200. doi: 10.1177/2211068211433503. Epub 2012 Feb 14 .

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