Proof of Concept for a Portable Platform for Molecular Diagnosis of Tropical Diseases: On-Chip Ready-to-Use Real-Time Quantitative PCR for Detection of Trypanosoma cruzi or Plasmodium spp.

Although molecular diagnostics is well established in clinical laboratories, its full potential has not been extended to field settings. Typically, diagnostic real-time quantitative PCR (qPCR) reagents require temperature-controlled transportation and storage. Furthermore, thermocyclers are bulky and fragile, requiring good infrastructure for optimal operation. These major hurdles strongly limit use of molecular-based tests in low-resource scenarios. Herein, Trypanosoma cruzi or Plasmodium spp. DNA were detected with qPCR using commercial equipment (ABI7500 instrument) and a prototype platform comprising a portable device and a silicon chip, named Q3-Plus. In addition, a ready-to-use reaction format, where all qPCR reagents are stored on plate or on chip, was compared with the traditional freezer-stored format. No significant differences were observed in detecting T. cruzi or Plasmodium spp. DNA between thermocyclers, as well as between reagents' formats, for storage periods of up to 28 days (at 2°C to 8°C or 21°C to 23°C, respectively). When challenged with patients' samples, the Q3-Plus system performed as efficiently as the standard equipment for Plasmodium spp. DNA detection, showing it to be a valuable solution to malaria point-of-care diagnostics. Detection of T. cruzi DNA in chronic patients' samples using the Q3-Plus system yielded approximately 50% efficiency relative to the ABI7500. These results are essential to support future endeavors to bring molecular diagnostics to the point of care, where most needed.

Key Enabling Technologies for Point-of-Care Diagnostics.

A major trend in biomedical engineering is the development of reliable, self-contained point-of-care (POC) devices for diagnostics and in-field assays. The new generation of such platforms increasingly addresses the clinical and environmental needs. Moreover, they are becoming more and more integrated with everyday objects, such as smartphones, and their spread among unskilled common people, has the power to improve the quality of life, both in the developed world and in low-resource settings. The future success of these tools will depend on the integration of the relevant key enabling technologies on an industrial scale (microfluidics with microelectronics, highly sensitive detection methods and low-cost materials for easy-to-use tools). Here, recent advances and perspectives will be reviewed across the large spectrum of their applications.

Q3: A Compact Device for Quick, High Precision qPCR.

An accurate and easy-to-use Q3 system for on-chip quantitative real-time Polymerase Chain Reaction (qPCR) is hereby demonstrated, and described in detail. The qPCR reactions take place inside a single-use Lab-on-a-Chip with multiple wells, each with 5 to 15 µL capacity. The same chip hosts a printed metal heater coupled with a calibrated sensor, for rapid and accurate temperature control inside the reaction mixture. The rest of the system is non-disposable and encased in a 7 × 14 × 8.5 (height) cm plastic shell weighing 300 g. Included in the non-disposable part is a fluorescence read-out system featuring up to four channels and a self-contained control and data storage system, interfacing with an external user-friendly software suite. Hereby, we illustrate the engineering details of the Q3 system and benchmark it with seamlessly ported testing protocols, showing that Q3 equals the performance of standard commercial systems. Overall, to the best of our knowledge, this is one of the most mature general-purpose systems for on-chip qPCR currently available.