Contents lists available at ScienceDirect Biosensors and Bioelectronics journal homepage: www.elsevier.com/locate/bios Smartphone-based mobile digital PCR device for DNA quantitative analysis with high accuracy Tong Gou a,1 , Jiumei Hu a,b,1 , Wenshuai Wu a,b , Xiong Ding a,b , Shufang Zhou a , Weibo Fang a , Ying Mu a,b, ⁎ a Research Center for Analytical Instrumentation, Institute of Cyber-Systems and Control, State Key Laboratory of Industrial Control Technology, Zhejiang University, Hangzhou 310027, PR China b College of Life Sciences, Zhejiang University, Hangzhou 310058, Zhejiang, PR China ARTICLE INFO Keywords: Digital PCR Smartphone Microfluidic chip Resource-limited settings Point-of-care Cancer biomarker gene ABSTRACT Digital polymerase chain reaction (dPCR) circumventing the external calibration and potentially providing absolute quantification of nucleic acids has become an increasingly popular manifestation of PCR in biological researches. However, currently reported or commercial dPCR devices are not suitable for applications in la- boratories or zones with limited infrastructures, due to low function integration, cost-inefficiency, or weak mobility. Herein, in order to enable accurate DNA quantitative analysis in such situations, we have developed a smartphone-based mobile dPCR device integrated with thermal cycling control, on-chip dPCR, data acquisition, and result analysis. All the function units are automatically controlled using a customized Android software. The device is approximately 90 mm × 90 mm × 100 mm in size and about 500 g in weight, only costing about 320 dollars except the smartphone. Coupled with the self-priming dPCR chip previously developed by our lab, the device is able to accurately quantify down to 10 copies of the human 18 S ribosomal DNA fragment inserted in a plasmid. Comparing to the commercial QuantStudio™ 3D dPCR platform, our device achieves a comparable analytical accuracy. Besides, our device is capable of detecting single molecule of cancer biomarker gene CD147 in a low number of HepG2 cells. Therefore, our dPCR device as a low-cost, potable, and robust tool for highly accurate DNA quantitative analysis has a great potential in Point-of-care (POC) applications. 1. Introduction The analysis of nucleic acids plays an important role in a variety of applications such as pathogen diagnostics of infectious diseases (Stramer et al., 2004, 2011), bioprospecting (Balcazar et al., 2015; Hafez et al., 2014), public health surveillance (Ecker et al., 2009; Quick et al., 2016), food safety supervision (DebRoy et al., 2011; Niessen et al., 2013), and environmental conservation (Piaggio et al., 2014; Thomsen et al., 2012). Currently, polymerase chain reaction (PCR) (Mullis et al., 1986) is the mainstay for nucleic acid analysis, due to the capability of amplifying small amounts of nucleic acid molecules from biological samples. In traditional PCR, the detection of amplified products greatly de- pends on gel electrophoresis assays, which is time-consuming and hard to quantify the target molecules (Hein et al., 2006). Therefore, real-time quantitative PCR (qPCR) is developed to readily quantify the products (Heid et al., 1996). However, the accuracy of quantitation in qPCR especially for rare allele detection is influenced by amplification bias or inhibitors. To address this challenge, digital PCR (dPCR) emerges as a promising alternative to conventional qPCR, due to the capability of absolute quantification which avoids the establishment of calibration using DNA standards (Xiong and Ying, 2016). According to the strategies of partitioning the loaded samples, ex- isting dPCR techniques can be mainly grouped into microfluidic chip- based dPCR (cdPCR) and droplet-based dPCR (ddPCR). At present, commercial dPCR device platforms are available, including cdPCR- based Biomark™ HD system from Fluidigm, QuantStudio 3D cdPCR system from ThermoFisher, and ddPCR-based QX200™ system from Bio- Rad Laboratories. In research fields, some customized dPCR devices are also reported. Using multi-material 3D printing, Begolo et al. designed a pumping lid-integrated SlipChip cdPCR device to achieve equipment- free pumping for reagent loading (Begolo et al., 2014). Selck and https://doi.org/10.1016/j.bios.2018.08.030 Received 29 May 2018; Received in revised form 12 August 2018; Accepted 13 August 2018 ⁎ Corresponding author at: Research Center for Analytical Instrumentation, Institute of Cyber-Systems and Control, State Key Laboratory of Industrial Control Technology, Zhejiang University, Hangzhou 310027, PR China. 1 These authors contributed equally to this work. E-mail address: muying@zju.edu.cn (Y. Mu). Biosensors and Bioelectronics 120 (2018) 144–152 Available online 17 August 2018 0956-5663/ © 2018 Elsevier B.V. All rights reserved. T