Development of Capillary Loop Convective Polymerase Chain Reaction Platform with Real-Time Fluorescence Detection

Abstract

Polymerase chain reaction (PCR) has been one of the principal techniques of molecular biology and diagnosis for decades. Conventional PCR platforms, which work by rapidly heating and cooling the whole vessel, need complicated hardware designs, and cause energy waste and high cost. On the other hand, partial heating on the various locations of vessels to induce convective solution flows by buoyancy have been used for DNA amplification in recent years. In this research, we develop a new convective PCR platform, capillary loop convective polymerase chain reaction (clcPCR), which can generate one direction flow and make the PCR reaction more stable. The U-shaped loop capillaries with 1.6 mm inner diameter are designed as PCR reagent containers. The clcPCR platform utilizes one isothermal heater for heating the bottom of the loop capillary and a CCD device for detecting real-time amplifying fluorescence signals. The stable flow was generated in the U-shaped container and the amplification process could be finished in 25 min. Our experiments with different initial concentrations of DNA templates demonstrate that clcPCR can be applied for precise quantification. Multiple sample testing and real-time quantification will be achieved in future studies.

A microfluidic system for rapid nucleic acid analysis based on real-time convective PCR at point-of-care testing

“…When the CPCR reactor is properly heated with one or two constant temperatures, a continuous circulatory flow will be developed to spontaneously and repeatedly transport the reagent through different temperature zones for amplification (Rajendran et al 2019a , b ). Comparing to conventional PCR, nucleic acid amplification with CPCR can be performed within remarkably short time with pseudo-isothermal heating (Li et al 2016 ; Chou et al 2017 ; Shu et al 2017 ).…”