VEGF Detection via Simplified FLISA Using a 3D Microfluidic Disk Platform

Abstract

Fluorescence-linked immunosorbent assay (FLISA) is a commonly used, quantitative technique for detecting biochemical changes based on antigen–antibody binding reactions using a well-plate platform. As the manufacturing technology of microfluidic system evolves, FLISA can be implemented onto microfluidic disk platforms which allows the detection of trace biochemical reactions with high resolutions. Herein, we propose a novel microfluidic system comprising a disk with a three-dimensional incubation chamber, which can reduce the amount of the reagents to 1/10 and the required time for the entire process to less than an hour. The incubation process achieves an antigen–antibody binding reaction as well as the binding of fluorogenic substrates to target proteins. The FLISA protocol in the 3D incubation chamber necessitates performing the antibody-conjugated microbeads’ movement during each step in order to ensure sufficient binding reactions. Vascular endothelial growth factor as concentration with ng mL−1 is detected sequentially using a benchtop process employing this 3D microfluidic disk. The 3D microfluidic disk works without requiring manual intervention or additional procedures for liquid control. During the incubation process, microbead movement is controlled by centrifugal force from the rotating disk and the sedimentation by gravitational force at the tilted floor of the chamber.

Keywords

• fluorescence-linked immunosorbent assay
• lab-on-a-disk
• vascular endothelial growth factor
• 3D microstructure