TRANSCRIPTOME 2002: From Functional Genomics to Systems
Poster PDF file
Affinity-Based Immobilization Tools for Functional Genomics
Amy L. Springer, Anna S. Gall, Karin A. Hughes, Robert J. Kaiser, Guisheng Li, Deborah D. Lucas, Kevin P. Lund, Doug A. Spicer, Jean P. Wiley, Prolinx, Inc., Bothell, WA
Studies of gene expression and subsequent interactions of gene products on a systems scale require methods that are robust, simple to implement and yield data that can be analyzed efficiently. Such methods often require immobilization of biomolecules for capture and detection of analytes, impurities, secondary products or metabolites. Immobilization methods such as direct conjugation to surfaces (e.g., glutaraldehyde coupling) or biologically-based affinity systems (e.g., (strept) avidin/biotin), can be limited by poor reproducibility, low surface capacities, essential purification steps and significant non-specific binding. Prolinx®, Inc. has developed a small molecule affinity system suitable for immobilization of nucleic acids and proteins on a variety of surfaces. This technology is based on the reversible complexation of phenyl(di)boronic acid (P(D)BA) with salicylhydroxamic acid (SHA). Surfaces can be reproducibly modified with SHA resulting in high capacities for P(D)BA-modified biomolecules and excellent assay sensitivities. P(D)BA-modification is performed in solution, independent of immobilization, and PDBA-conjugates can be directly immobilized on an SHA-modified solid support without purification; any excess reagent is removed by washing. As a result, multiple conjugations may be performed in an automated format suitable for high-throughput applications such as protein microarrays. The advantages demonstrated using this system make P(D)BA-SHA technology a convenient platform for systems-scale research.
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