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Published on: November 13, 2021
A flow-based enzyme-linked immunosorbent assay on a polydimethylsiloxane microchip for the rapid determination of
Hizuru Nakajima1, Maiko Yagi, Yuki Kudo
1Faculty of Urban Environmental Sciences, Tokyo Metropolitan University, 1-1 Minamiohsawa, Hachioji, Tokyo 192-0397, Japan.
Insights
A novel flow-based enzyme-linked immunosorbent assay (ELISA) on a microchip significantly speeds up immunoglobulin A (IgA) detection. This method drastically reduces reaction times and reagent usage compared to traditional assays.
Area of Science:
- Biotechnology
- Analytical Chemistry
- Microfluidics
Background:
- Conventional enzyme-linked immunosorbent assays (ELISA) are widely used for detecting specific analytes like immunoglobulin A (IgA).
- Traditional ELISA methods often require lengthy incubation and reaction times, limiting their throughput and efficiency.
- The development of microfluidic platforms offers potential for miniaturization and faster assay formats.
Purpose of the Study:
- To develop a rapid, flow-based enzyme-linked immunosorbent assay (ELISA) for immunoglobulin A (IgA) determination.
- To miniaturize the ELISA process using a polydimethylsiloxane (PDMS) microchip.
- To significantly reduce assay time and reagent consumption compared to conventional methods.
Main Methods:
- A polydimethylsiloxane (PDMS) microchip was fabricated for flow-based assays.
- A sandwich-type ELISA principle was employed, immobilizing primary antibodies (anti-IgA) in the microchannel.
- Detection of immunoglobulin A (IgA) utilized a secondary antibody labeled with horseradish peroxidase (HRP) and Amplex Red substrate with fluorescent imaging.
Main Results:
- The flow-based microchip ELISA demonstrated a linear calibration curve for IgA standards from 0-50 ng/mL at a flow rate of 10 µL/min.
- The reaction time was dramatically reduced to 4.8 seconds, a 375-fold decrease compared to the 30-minute reaction time of conventional assays.
- Reagent and sample volumes were reduced by approximately 100-fold.
Conclusions:
- A rapid and efficient flow-based ELISA on a PDMS microchip has been successfully developed for IgA detection.
- This microfluidic approach offers significant advantages in terms of speed and reduced reagent/sample consumption.
- The developed method holds promise for faster and more economical diagnostic applications.
Abstract:
A flow-based enzyme-linked immunosorbent assay (ELISA) on a polydimethylsiloxane (PDMS) microchip has been developed for the rapid determination of immunoglobulin A (IgA). The analytical principle of this integrated method is the same as the conventional sandwich-type ELISA. A primary antibody (anti-IgA) was adsorbed on the surface of a PDMS microchannel, and then an antigen (IgA) and a secondary antibody (anti-IgA HRP labeled) were reacted successively. The resulting antigen-antibody complex, fixed on the surface of the microchannel, was detected using Amplex((R)) Red and a fluorescent imaging system. The calibration curve of the IgA standard solution was linear in the range of 0-50ng/mL at the flow rate of 10muL/min. This flow rate corresponds to the reaction time of 4.8s. Compared to the conventional assay on a 96-well microtiter plate, the present assay on the microchip dramatically shortened the reaction time necessary for the enzyme-substrate reaction from 30min to 4.8s, i.e., to 1/375. The amounts of the reagent and sample were also reduced to 1/100 compared to the 96-well microtiter plate.
