Paper-Based Radial Flow Assay Integrated to Portable Isothermal Amplification Chip Platform for Colorimetric...

Area of Science:

• Biotechnology
• Analytical Chemistry
• Materials Science

Background:

• Developing sensitive and portable methods for on-site DNA detection is crucial for various applications, including diagnostics and environmental monitoring.
• Existing methods often require complex equipment and trained personnel, limiting their use in resource-limited settings.
• Integrated microfluidic systems offer a promising platform for simplifying and miniaturizing molecular detection assays.

Purpose of the Study:

• To develop a novel integrated detection system combining paper-chip technology with a polydimethylsiloxane (PDMS) microchip for on-site colorimetric DNA detection.
• To optimize padlock probe DNA (PLP)-mediated rolling circle amplification (RCA) and radial flow assay for sensitive and specific DNA detection.
• To evaluate the system's performance, including detection limit, specificity, and suitability for resource-limited environments.

Main Methods:

• An integrated system was designed using a PDMS microchip and a paper-based detection strategy.
• Padlock probe DNA (PLP)-mediated rolling circle amplification (RCA) was employed for signal amplification.
• A radial flow assay with Au-probe labeling was utilized for colorimetric visualization of DNA.
• Ligation, RCA, and labeling reactions were performed in a single chamber under isothermal conditions within the PDMS chip.
• The reaction solution was then transferred to a paper chip for signal readout.

Main Results:

• The integrated system achieved a DNA detection limit of 14.7 nM.
• The system demonstrated high specificity, successfully discriminating target DNA from sequences with single-base mismatches.
• Optimization of the PDMS chip's chamber design and temperature control system contributed to an optimal analysis environment.
• The developed system is cost-effective due to the use of inexpensive paper and PDMS materials.

Conclusions:

• The novel integrated detection system provides a sensitive, specific, and user-friendly platform for on-site colorimetric DNA detection.
• The system is suitable for applications such as single nucleotide polymorphism identification and pathogen gene detection.
• Its portability, cost-effectiveness, and minimal equipment requirements make it ideal for resource-limited settings.