Parallel microfluidic surface plasmon resonance imaging arrays.
Surface plasmon resonance imaging (SPRi) is a label-free technique used for the quantitation of binding affinities and concentrations for a wide variety of target molecules. Although SPRi is capable of determining binding constants for multiple ligands in parallel, current commercial instruments are limited to a single analyte stream on multiple ligand spots. Measurement of binding kinetics requires the serial introduction of different analyte concentrations; such repeated experiments are conducted manually and are therefore time-intensive. To address these challenges, we have developed an integrated microfluidic array using soft lithography techniques for high-throughput SPRi-based detection and determination of binding affinities of antibodies against protein targets. The device consists of 264 element-addressable chambers isolated by microvalves. The resulting 700 pL chamber volumes, combined with a serial dilution network for simultaneous interrogation of up to six different analyte concentrations, allow for further speeding detection times. To test for device performance, human alpha-thrombin was immobilized on the sensor surface and anti-human alpha-thrombin IgG was injected across the surface at different concentrations. The equilibrium dissociation constant was determined to be 5.0 +/- 1.9 nM, which agrees well with values reported in the literature. The interrogation of multiple ligands to multiple analytes in a single device was also investigated and samples were recovered with no cross-contamination. Since each chamber can be addressed independently, this array is capable of interrogating binding events from up to 264 different immobilized ligands against multiple analytes in a single experiment. The development of high-throughput protein analytic measurements is a critical technology for systems approaches to biology and medicine.
Ouellet
Eric
E
Michael Smith Laboratories, University of British Columbia, Vancouver, BC, Canada V6T 1Z4.
Lausted
Christopher
C
Lin
Tao
T
Yang
Cheng Wei T
CW
Hood
Leroy
L
Lagally
Eric T
ET
D004867:Equipment Design; D019544:Equipment Failure Analysis; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D040081:Protein Array Analysis / Q000295:instrumentation*; D025941:Protein Interaction Mapping / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance / Q000295:instrumentation*