Multiplexed, rapid, point of care device to quantify allergen-specific IgE

In this paper, we have integrated our in-house Interferometric Reflectance Imaging Sensor (IRIS), a simple, sensitive, and cost-effective technology for label-free detection of proteins in multiplexed format, with enhanced fluorescence technology into a calibrated, compact, inexpensive, and multiplexed point-of-care protein microarray sensor called the Calibrated Fluorescence Enhancement (CaFE) platform. This platform utilizes both label-free and fluorescence modalities in order to correlate surface probe density to captured labeled secondary antibody. The platform includes a chip fabricated with two areas of oxide layers with different thicknesses: one area with a 500 nm SiO₂ layer optimized for interferometric label-free detection and quantification of proteins and one area with a 100 nm SiO₂ layer optimized for enhanced fluorescence over a broad range of fluorophores that are commonly used in bioassays. Probes spotted on the thick region are measured to quantify the capturing immobilized probe density on the platform. However, given a specific fluorophore, we can design a chip of a single thickness that operates well at both label-free and enhanced fluorescence measurements for single spot analysis. We have identified three major technological advantages of our Calibrated Enhancement (CaFE) platform: a ten-fold gain in fluorescence signal due to optimized substrate; a calibrated, quantitative method that uses the combined fluorescent and label-free modalities to accurately measure the density of probe and bound target for a variety of systems; and a compact measurement platform offering reliable and rapid results at the doctor´s office for any test that uses a known antibody-antigen pair.