University of Washington

Kjell E Nelson

Paul Yager

Turgut Fettah Kosar

Paolo Spicar-Mihalic

Dean Stevens

Michael Wai-Haung Look

Rahber Thariani

Afshin Mashadi-Hossein

Katherine McKenzie

Patrick S Stayton

Elain S Fu

Melissa S Hasenbank

Todd C McDevitt

Disclosed are methods and devices for rapid parallel molecular affinity assays performed in a microfluidic environment. The invention exploits hydrodynamic addressing to provide simultaneous performance of multiple assays in parallel using a minimal sample volume flowing through a single channel.

Method and device for rapid parallel microfluidic molecular affinity assays

Disclosed is a microfluidic assay system and methods that apply flow injection analysis to permit dispersion monitoring. A solution containing a reagent that binds an analyte and a tracer is delivered via pressure-driven flow into the receiving end of the injection channel of the system of the invention. A sample fluid suspected of containing the analyte is delivered into the upstream end of the input channel under conditions permitting flow of the sample fluid toward the downstream end of the assay channel and permitting dispersion of the reagent into the sample fluid. The amount of tracer present in the fluid as it passes over the reference region and the capture region and the amount of binding between the analyte and the capture region are detected. The amount of binding detected between the analyte and the capture region is correlated to the amount of tracer detected in the reference region.

Microfluidic assay system with dispersion monitoring

The present invention provides methods and compositions for amplifying the detection signal in surface plasmon resonance (SPR)-based flow systems. The signal amplification methods comprise the use of well established marker systems that provide a precipitate. The marker systems include, for example, enzyme and nucleation systems. Enzymes suitable for use as a marker system include peroxidases and phosphatases. The amplification system is useful in any SPR-based detection system including microfluidic systems, e.g., 'lab on a chip' systems and the like. The methods can comprise any SPR-based assay format, including typical immunoassay formats. The immunoassay formats can include competitive and sandwich assays. Analyte capture agents can include antibodies, lectins, carbohydrates, polynucleotides, receptor proteins, and the like.

Signal amplification method for surface plasmon resonance-based chemical detection

Streptavidin molecules are disclosed that contain a secondary functional domain. In preferred embodiments, the secondary domain is a cell adhesion peptide incorporated in the streptavidin amino acid sequence at a site not interfering with biotin binding. In a preferred embodiment, the cell adhesion peptide is arginine-glycine-aspartate (Arg-Gly-Asp) (RGD). The peptide is preferably placed on an exposed loop of the streptavidin molecule, such as within the loop defined by residues 63 to 69. The mutant streptavidin molecule can have other characteristics such as reduced biotin binding due to a modification of an amino acid at the biotin binding site. Preferred uses for the disclosed streptavidin molecules are as adaptors to bring, via a streptavidin/biotin interaction, the secondary functional domain into proximity with a cell or molecule to be affected and as a coating for substrates such as vascular devices or prostheses.

Streptavidin mutants having secondary functional domains

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Seattle, WA, United States of America

Kjell E Nelson