On-chip RT-LAMP and colorimetric detection of the prostate cancer 3 biomarker with an integrated thermal and imaging box

Abstract

To achieve low-cost, compact, and portable nucleic acid testing, an integrated device containing a three-dimensional printing fabricated reverse transcription loop-mediated isothermal amplification (RT-LAMP) chip, a thermal module, and an imaging module was developed. Samples and RT-LAMP reagents were loaded on a sponge-like polyvinyl alcohol pad on a chip, whereas the colorimetric detection zone was a dry paper pre-loaded with Calcine. The sealed chip was incubated on the integrated thermal module, and the RT-LAMP products were pressed into the Calcine pre-loaded dry paper by a stick. Colorimetric changes could be visually observed by the naked eye or imaged with a smartphone camera through the imaging module. For detection of the prostate cancer antigen 3 (PCA3) biomarker, LAMP primers were designed and verified. The specificity of Calcine pre-loaded dry paper based on colorimetric detection of positive LAMP products was investigated. The reaction conditions for on-chip RT-LAMP such as amplification time, temperature, and volume were optimized. Finally, a detection limit of 0.34 fg/μL RNA was achieved with the proposed on-chip RT-LAMP and colorimetric detection method for PCA3. Since the thermal plate is powered by a 12-V battery and the color change can be imaged with a smartphone, the integrated platform can be operated on-site, highlighting its potential in point-of-care testing applications.

Keywords: Colorimetric; Microfluidic; Paper based analytical device; RT-LAMP; Smartphone.