Picoliter Well Array Chip-Based Digital Recombinase Polymerase Amplification for Absolute Quantification of Nucleic Acids

Li, Zhao; Liu, Yong; Wei, Qingquan; Liu, Yuanjie; Liu, Wenwen; Zhang, Xuelian; Yu, Yude

ImageJ (NIH, Bethesda, MD, USA) and custom Matlab code were used to systematically detect and quantify fluorescent "positive" microwells and to analyze their density and averaged fluorescence intensity [35] . Background subtractions and contrast enhancement were performed for quantification of the dRPA results. The image acquired at the beginning (0 min) was regarded as the background noise of the experiment; other images acquired subsequently subtract it in situ for contrast enhancement. To enhance the visual contrast, grayscale images were converted to green. The quantitative results were then compared to the expected number of positive microwells predicted from Poisson statistics for serial dilutions of the known sample concentration.