4. Conclusions
The aim of this study was to fabricate biocompatible, low-cost, and disposable chips with integrated heater, which should be able to perform DNA amplification, and possible in situ fluorescence detection in the near future. In this case there is no interest in quantification of the DNA, but only in amplification of DNA until the detection threshold is reached. As proof-of-principle the MDA reaction and ex-situ fluorescence measurements were used.
With the proposed fabrication process, low-cost and biocompatible chips (Figure 12b) were fabricated. The integrated resistive heaters on the chips were characterized and showed a temperature stability of ±2 °C over a time period of 25 h, which is at least twelve-fold longer than the required operating times for DNA amplification reactions [6,8,9,10,11]. The main cause of this period of lowered temperature was due to the fact that the measurement was run overnight.
With the proof-of-principle device, successful DNA amplifications using MDA inside a disposable polymeric chip were achieved. The heat for the reaction was applied using the integrated low-cost Au-resistive heater. The device was operated at a suitable temperature for MDA reactions and the amplified DNA was measured using EvaGreen fluorescence dye and an ex situ spectrofluorometer. A distinct peak is visible in the reaction mixtures which is absent in the NTC mixtures. The operating temperature for MDA reactions is around 30 °C, which is comparable with a nice summer day. Using amplification reactions which such low reaction temperatures could encounter problems at warmer locations. However, as MDA is not sequence specific, this reaction will not be integrated in the final protocols. MDA was only used as proof-of-principle reaction to show the biocompatibility of the device and functioning of the integrated heater. Sequence specific amplifications, e.g., HDA and LAMP, are performed at higher temperatures, as will be discussed in Section 5. This makes the system less sensitive to the hot summer days.
The device in its current state is not fully conform the WHO-SDI ASSURED criteria [49] as it still relies on the use of (expensive) external equipment. However, the first steps are made to an ASSURED device. Future steps which will make the device fully ASSURED are given in the next section (Section 5).