High-speed interferometric detection of label-free immunoassays on the biological compact disc

Background: We describe a direct-detection immunoassay that uses high-speed optical interferometry on a biological compact disc (BioCD). Methods: We fabricated phase-contrast BioCDs from 100-mm diameter 1.1-mm thick borosilicate glass disks coated with a 10-layer dielectric stack of Ta₂O ₅/SiO₂ that serves as a mirror with a center wavelength at 635 nm. The final layer is a λ/4 layer of SiO₂ onto which protein patterns are immobilized through several different chemical approaches. Protein on the disc is scanned by a focused laser spot as the disc spins. Interaction of the light with the protein provides both a phase-modulated signal and a local reference that are combined interferometrically to convert phase into intensity. A periodic pattern of protein on the spinning disc produces an intensity modulation as a function of time that is proportional to the surface-bound mass. The binding of antigen or antibodies is detected directly, without labels, by a change in the interferometric intensity. The technique is demonstrated with a reverse assay of immobilized rabbit and mouse IgG antigen incubated against anti-IgG antibody in a casein buffer. Results: The signal increased with increased concentration of analyte. The current embodiment detected a concentration of 100 ng/L when averaged over ∼3000 100-micron-diameter protein spots. Conclusions: High-speed interferometric detection of label-free protein assays on a rapidly spinning BioCD is a high-sensitivity approach that is amenable to scaling up to many analytes.