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Proceedings Paper

Real-time spinning-disk interferometric immunoassays
Author(s): Manoj M. Varma; Halina D. Inerowicz; Fred E. Regnier; David D. Nolte
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Paper Abstract

We describe real-time detection of antigen/antibody binding using a high-speed label-free interferometric detection technique. The sensor, called the BioCD, consists of microfabricated gold interferometric structures on 2” dielectric laser mirror substrates that spin at rates up to 6000 rpm. The interferometric microdiffraction elements operate in the linear sensitivity regime of the interferometer. Antibodies or proteins are immobilized on the gold interferometric structures through an intermediate thiol layer. The molecules are immobilized by application of reagents or samples to the disk while it is spinning. The centrifugal force distributes the sample over the sensor surface, causing a change in the optical phase of the interferometric element, which is detected in real-time using lock-in detection with small detection bandwidth. The thiolated BioCD is spun at 1500 rpm and anti-mouse IgG, rabbit IgG and mouse IgG are delivered in succession to the sensor surface with potash buffered saline (PBS) wash cycles interspersed between each exposure to remove excess unbound proteins. The layer of anti-mouse IgG binds to the thiolated gold sensor elements which later bind specific mouse IgG. We have observed a 10% change in the interferometric signal when mouse IgG binds to an immobilized layer of anti-mouse IgG. No significant non-specific binding of rabbit IgG was detected. The sensitivity and throughput of this sensor will be discussed. An advantage of this new approach, relative to previous work in which the disk was incubated with antigen off-line, is the real-time detection of antigen binding, which could be valuable for simultaneous high-speed screening of a large number of protein interactions.

Paper Details

Date Published: 22 June 2004
PDF: 7 pages
Proc. SPIE 5328, Microarrays and Combinatorial Techniques: Design, Fabrication, and Analysis II, (22 June 2004); doi: 10.1117/12.529531
Show Author Affiliations
Manoj M. Varma, Purdue Univ. (United States)
Halina D. Inerowicz, Purdue Univ. (United States)
Fred E. Regnier, Purdue Univ. (United States)
David D. Nolte, Purdue Univ. (United States)

Published in SPIE Proceedings Vol. 5328:
Microarrays and Combinatorial Techniques: Design, Fabrication, and Analysis II
Dan V. Nicolau; Ramesh Raghavachari, Editor(s)

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