Real-time capillary convective PCR based on horizontal thermal convection

Abstract

A real-time, capillary, convective polymerase chain reaction (PCR) system based on horizontal convection is developed and analyzed. The capillary tube reactor is heated at one end in a pseudo-isothermal manner to achieve efficient thermal cycling based on horizontal thermal convection. Mathematical modeling and the in silico simulations indicate that, once consistent temperature gradient along the horizontal capillary tube has been established, a repeatable and continuous circulatory flow in the horizontal directions is created. The formed convection is able to transport PCR reagents through different temperature zones inside the horizontal capillary tube for different reaction stages, that is, DNA denaturing, annealing, and extension in a typical PCR cycle. Furthermore, the effectiveness and efficiency of the horizontal convection for PCR thermal cycling in a capillary tube is confirmed by experimentation. To evaluate the concept of horizontal convective PCR, a compact system, which is able to heat the capillary tube from one end and monitor the fluorescence in situ with a smartphone camera, is developed for real-time amplification. With horizontal thermal convection, influenza A (H1N1) virus nucleic acid targets with a limit of detection (LOD) of 1.0 TCID50/mL can be successfully amplified and detected in 30 min, which is promising for efficient nucleic acid analysis in point-of-care testing.