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Proceedings Paper

Affinity reagent technology development and application to rapid immunochromatographic pathogen detection
Author(s): Letha J. Sooter; Dimitra N. Stratis-Cullum; Yanting Zhang; Patrick S. Daugherty; H. Tom Soh; Paul Pellegrino; Nancy Stagliano
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Paper Abstract

Immunochromatography is a rapid, reliable, and cost effective method of detecting biowarfare agents. The format is similar to that of an over-the-counter pregnancy test. A sample is applied to one end of a cassette and then a control line, and possibly a sample line, are visualized at the other end of the cassette. The test is based upon a sandwich assay. For the control, a line of Protein A is immobilized on the membrane. Gold nanoparticle bound IgG flows through the membrane and binds the Protein A, creating a visible line on the membrane. For the sample, one epitope is immobilized on the membrane and another epitope is attached to gold nanoparticles. The sample binds gold bound epitope, travels through the membrane, and binds membrane bound epitope. The two epitopes are not cross-reactive, therefore a sample line is only visible if the sample is present. In order to efficiently screen for binders to a sample target, a novel, Continuous Magnetic Activated Cell Sorter (CMACS) has been developed on a disposable, microfluidic platform. The CMACS chip quickly sorts E. coli peptide libraries for target binders with high affinity. Peptide libraries, are composed of approximately ten million bacteria, each displaying a different peptide on their surface. The target of interest is conjugated to a micrometer sized magnetic particle. After the library and the target are incubated together to allow binding, the mixture is applied to the CMACS chip. In the presence of patterned nickel and an external magnet, separation occurs of the bead-bound bacteria from the bulk material. The bead fraction is added to bacterial growth media where any attached E. coli grow and divide. These cells are cloned, sequenced, and the peptides are assayed for target binding affinity. As a proof-of-principle, assays were developed for human C-reactive protein. More defense relevant targets are currently being pursued.

Paper Details

Date Published: 5 October 2007
PDF: 8 pages
Proc. SPIE 6759, Smart Biomedical and Physiological Sensor Technology V, 67590A (5 October 2007); doi: 10.1117/12.732731
Show Author Affiliations
Letha J. Sooter, Army Research Lab. (United States)
Dimitra N. Stratis-Cullum, Army Research Lab. (United States)
Yanting Zhang, CytomX, LLC (United States)
Patrick S. Daugherty, Univ. of California/Santa Barbara (United States)
H. Tom Soh, Univ. of California/Santa Barbara (United States)
Paul Pellegrino, Army Research Lab. (United States)
Nancy Stagliano, CytomX, LLC (United States)


Published in SPIE Proceedings Vol. 6759:
Smart Biomedical and Physiological Sensor Technology V
Brian M. Cullum; D. Marshall Porterfield, Editor(s)

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