Low-cost, real-time, continuous flow PCR system for pathogen detection

Abstract

In this paper, we present a portable and low cost point-of-care (POC) PCR system for quantitative detection of pathogens. Our system is based on continuous flow PCR which maintains fixed temperatures zones and pushes the PCR solution between two heated areas allowing for faster heat transfer and as a result, a faster PCR. The PCR system is built around a 46.0 mm × 30.9 mm × 0.4 mm disposable thermoplastic chip. In order to make the single-use chip economically viable, it was manufactured by hot embossing and was designed to be compatible with roll-to-roll embossing for large scale production. The prototype instrumentation surrounding the chip includes two heaters, thermal sensors, and an optical system. The optical system allows for pathogen detection via real time fluorescence measurements. FAM probes were used as fluorescent reporters of the amplicons generated during the PCR. To demonstrate the function of the chip, two infectious bacteria targets were selected: Chlamydia trachomatis and Escherichia coli O157:H7. For both bacteria, the limit of detection of the system was determined, PCR efficiencies were calculated, and different flow velocities were tested. We have demonstrated successful detection for these two bacterial pathogens highlighting the versatility and broad utility of our portable, low-cost, and rapid PCR diagnostic device.

Tunable and precise miniature lithium heater for point-of-care applications

“…To the 4-mm 2 heater, 1.5 mg of SDS and 28.5 mg of mannitol were added; while the 1.5-mm 2 heater had 30 mg of SDS. The 4-mm 2 heater was first placed in 3 mL of water and 10% antifoam. The lithium was allowed to be fully consumed, after which the 1.5-mm 2 heater was placed in the solution to maintain the temperature at 42°C.…”

“…chemical heater | diagnostics | point of care P oint-of-care diagnostic assays often involve complex multistep reactions that require a wide variety of temperatures for steps ranging from sample processing to genetic analysis (1)(2)(3). Although precise heating is critical to running these assays, it is challenging to provide it in field settings away from established infrastructure.…”

An integrated biosensor platform for extraction and detection of nucleic acids

“…In this scenario, the integration of silicon technologies such as micro‐electromechanical systems and very large‐scale integration, together with microfluidics, is very intriguing to achieve full integration of NA testing protocols on miniaturized and portable diagnostic devices (Fernández‐Carballo et al, 2016; Guarnaccia et al, 2014; Hsieh et al, 2015; Tong et al, 2019). The main advantages of silicon are low heat capacity, good thermal conductivity, good biocompatibility, chemical derivatization, and the possibility to produce patterned structures, which increase the surface‐area ratio, improving the efficiency of NA extraction and detection (Leonardi et al, 2018; Petralia et al, 2015; Valli et al, 2006).…”

Analysis of PCR Kinetics inside a Microfluidic DNA Amplification System

“…In the unidirectional reactor, only an external device is needed to push the DNA sample through a single channel. The sample flows into the channel, experiences several temperature regions repeatedly, exits from the outlet, and completes the amplification process [ 5 ]. A self-propelled CFPCR in a microfluidic device that requires no external pump to drive the flow was also proposed.…”