A reagent-centred thermal control system driven by a cascade temperature control algorithm for high-speed PCR

Accelerating quantitative polymerase chain reaction (qPCR) without compromising analytical fidelity remains a significant challenge in molecular diagnostics, primarily due to the thermal lag between heating elements and reagents. Here, we report a high-speed qPCR platform that overcomes this limitation using a reagent-centric cascade control strategy. The system employs a planar PCB-based copper heater that functions as both a heating element and a temperature sensor, ensuring low-latency, sensor-efficient thermal feedback. To overcome this thermal delay, a virtual temperature sensor-derived from system identification-is used to estimate the real-time reagent temperature, which drives an outer-loop fuzzy PID controller with feedforward compensation. Inner cascaded loops stabilize the heater current and surface temperature. The system achieves reagent-phase average heating and cooling rates of 24.1 °C s^-1 and 19 °C s^-1, respectively. Furthermore, the reagent temperature is controlled with an accuracy of ±0.2 °C and an overshoot of less than 0.2 °C. A complete 45-cycle amplification is achieved in as little as 4.4 minutes. Crucially, this speed is attained without analytical compromise, demonstrating excellent quantitative accuracy (<i>R</i>² = 0.9965) and amplification efficiency (109.8%). The proposed reagent-centred control framework offers a scalable pathway for developing high-throughput PCR and molecular diagnostic instruments, supporting fast, accurate, and scalable nucleic acid testing.