The overall goal of this proposal is to develop a robust, sensitive and reliable label-free detector that can evaluate the real-time binding kinetics of 10,000 spot small molecule chemical microarrays against various single component or multi-component analytes. In 1991, we first reported the one-bead one-compound combinatorial library (OBOC) method which is highly efficient and we have successfully used it to identify ligands to a large number of biological targets. Although the OBOC library method is highly versatile, the amount of compounds contained in one single bead is small (100 pico-mol) and is inadequate for many solution phase assays. Last year, we reported on the development of an encoded """"""""one-aggregate one-compound"""""""" library method, in which the powerful split-synthesis method is used for the construction of the library, and compounds at l -10 mmol range can be efficiently prepared. Very recently, we reported on the development and application of a novel chemical microarray method, in which small organic molecules or short peptides are chemo-selectively ligated to a polymer (e.g. agarose or protein) and then printed on glass, plastic microscope slides or PVDF membranes. We envision that by combining our highly efficient """"""""one-aggregate one-compound"""""""" library method with this novel chemical microarray technique, we easily can print thousands of replicates of high density small molecule microarrays (10,000 spots/ slide) and use them to probe a variety of biological analytes such as serum, cell extracts or pure proteins. Our hypothesis is that by combining our novel combinatorial chemistry and microarray platforms with a highly sensitive and reliable 2-D label-free optical detector that can efficiently measure real-time binding kinetics, we will be able to rapidly and accurately study the binding kinetics of a large number of analytes (e.g., individual protein or complex analytes such as whole serum) against a large number of immobilized small molecules, peptides, oligonucleotides or proteins.
Specific aims of this proposal include the design and construction of a prototype detector that can measure real-time binding kinetics of analytes (dissolved in the mobile phase) against 100 - and 400-spot chemical microarrays. The next phase will be to develop a detector that can analyze 10,000 spot microarrays in one single run. Five 10,000 small molecule encoded bead-aggregate libraries will be prepared and printed on glass slide replicates as microarrays, and these microarrays will be analyzed by the optical detector to be developed in this proposed research. ? ?

Agency
National Institute of Health (NIH)
Institute
National Human Genome Research Institute (NHGRI)
Type
Research Project (R01)
Project #
5R01HG003827-02
Application #
7168830
Study Section
Special Emphasis Panel (ZEB1-OSR-C (O1))
Program Officer
Ozenberger, Bradley
Project Start
2006-01-17
Project End
2009-11-30
Budget Start
2006-12-01
Budget End
2007-11-30
Support Year
2
Fiscal Year
2007
Total Cost
$587,215
Indirect Cost
Name
University of California Davis
Department
Physics
Type
Schools of Arts and Sciences
DUNS #
047120084
City
Davis
State
CA
Country
United States
Zip Code
95618
Fei, Yiyan; Sun, Yung-Shin; Li, Yanhong et al. (2015) Characterization of Receptor Binding Profiles of Influenza A Viruses Using An Ellipsometry-Based Label-Free Glycan Microarray Assay Platform. Biomolecules 5:1480-98
Landry, J P; Malovichko, G; Zhu, X D (2015) High-Throughput Dose-Response Measurement Using a Label-Free Microarray-in-Microplate Assay Platform. Anal Chem 87:5640-8
Sun, Yung-Shin; Zhu, Xiangdong (2014) Real-time, label-free detection of biomolecular interactions in sandwich assays by the oblique-incidence reflectivity difference technique. Sensors (Basel) 14:23307-20
Landry, James P; Fei, Yiyan; Zhu, Xiangdong et al. (2013) Discovering small molecule ligands of vascular endothelial growth factor that block VEGF-KDR binding using label-free microarray-based assays. Assay Drug Dev Technol 11:326-32
Fei, Yiyan; Landry, James P; Li, Yanhong et al. (2013) An optics-based variable-temperature assay system for characterizing thermodynamics of biomolecular reactions on solid support. Rev Sci Instrum 84:114102
Landry, James P; Fei, Yiyan; Zhu, Xiangdong (2012) Simultaneous measurement of 10,000 protein-ligand affinity constants using microarray-based kinetic constant assays. Assay Drug Dev Technol 10:250-9
Landry, James P; Fei, Yiyan; Zhu, X D (2011) High Throughput, Label-free Screening Small Molecule Compound Libraries for Protein-Ligands using Combination of Small Molecule Microarrays and a Special Ellipsometry-based Optical Scanner. Int Drug Discov :8-13
Fei, Y Y; Schmidt, A; Bylund, G et al. (2011) Use of real-time, label-free analysis in revealing low-affinity binding to blood group antigens by Helicobacter pylori. Anal Chem 83:6336-41
Fei, Yiyan; Sun, Yung-Shin; Li, Yanhong et al. (2011) Fluorescent labeling agents change binding profiles of glycan-binding proteins. Mol Biosyst 7:3343-52
Lo, Kai-Yin; Sun, Yung-Shin; Landry, James P et al. (2011) Label-free detection of surface markers on stem cells by oblique-incidence reflectivity difference microscopy. Biotechniques 50:381-8

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