Assessing Aptamer-Analyte Binding Kinetics by Microfluidic Fluorescence Microscopy.

ABSTRACT

Kinetic measurement plays a crucial role in understanding aptamer binding mechanisms and identifying appropriate aptamers for clinical and research applications. Current techniques, while well established, generally require large sample volumes, bulky and expensive instruments operated by trained personnel, and are hence not readily accessible to resource-limited research laboratories. This paper presents a fluorescence microscopy-based microfluidic assay for measuring aptamer-analyte binding kinetics in a simple and cost-effective manner. Kinetic measurements are achieved by monitoring time-course fluorescence of fluorescently labeled aptamers as they bind to the targets trapped in a microfluidic chip. Fluorescence measurements are performed on a standard fluorescence microscope and are accessible to laboratories with only modest resources. Moreover, microfluidic technology allows efficient and cost-effective immobilization of small amounts of target molecules or live cells as well as flow-based manipulation of aptamers for the measurements. Kinetic measurements of aptamer binding to immunoglobulin E protein and CCRF-CEM cells have yielded results consistent with those obtained from established methods, demonstrating the potential utility of our method for exploring aptamer-target interactions and identifying aptamers that best suit specific given biomedical applications.