A cell-free paper-based biosensor dependent on allosteric transcription factors (aTFs) for on-site detection of harmful metals Hg2+ and Pb2+ in water.

Water quality monitoring requires a reliable and practical on-site detection method for heavy metal ions. Combining an in vitro transcription (IVT) technology with allosteric transcription factors (aTFs), we developed a cell-free paper-based biosensor for on-site detection of Hg2+ and Pb2+ in water. Suitable aTFs screened using surface plasmon resonance (SPR) were employed for building biosensors. ATFs could disassociate from DNA due to their specific affinity to metal ions, and fluorescent RNA was transcribed as a signal. The developed biosensor could quantitatively detect Hg2+ in a linear dynamic range of 0.5-500 nM and Pb2+ in a 1-250 nM range in a 1 h period. The LOD of the biosensor was 0.5 nM for Hg2+ and 0.1 nM for Pb2+. The recoveries ranged from 91.09% to 123.24% for actual water samples detection. Furthermore, freeze-drying was used to create a paper-based biosensor that could detect Hg2+ and Pb2+ simultaneously on-site. This research presents a useful technique for various heavy metal ion detections.