An on-chip valve-assisted microfluidic chip for quantitative and multiplexed detection of biomarkers

In this study, we developed a microfluidic chip integrated with on-chip valves for quantitative detection of biomarkers. The sandwich-structured microfluidic chip consists of a top fluidic layer embedded with a zigzag channel, a middle tinfoil layer pre-patterned with protein stripes, and a bottom substrate layer with two waste chambers. A set of customized on-chip valves was introduced for precise control of fluid flow. To improve the production volume and uniformity among chips, we selected an injection molding method for mass-production of the main components. We standardized the process of sealing chip layers and assembling the valve holder to improve the effectiveness of cooperation with the instrument. To demonstrate the versatility of the microfluidic chip for quantitative detection of biomarkers, we used it to automatically detect C-reactive proteins (CRP), interleukin-6 (IL-6), insulin, and osteocalcin with high sensitivity and excellent reproducibility inside a customized and portable instrument. We further conducted the automated chip-based chemiluminescence immunoassay to test CRP in serum samples for demonstration of potential clinical applications, assess the activity of capture antibodies of CRP (CRP-Ab(1)) on the tinfoil layer for preliminary assessment of storage stability, and simultaneously detect CRP and IL-6 for confirmation of scalable multiplexation. This microfluidic chip for quantitative, automated, and portable immunoassay provides a promising choice for clinical practices.