Optimizing the performance of digital loop-mediated isothermal amplification

Digital loop-mediated isothermal amplification (dLAMP) is attractive for the detection of nucleic acid due to its superior characteristics including isothermal amplification, absolute quantification, and single-molecule sensitivity. However, dLAMP suffers from the inaccurate quantification caused by low digital efficiency, which means only part of loaded template molecules could be amplified. We here developed a prehybridization-induced enhancement (PIE) strategy which could improve digital efficiency about 2–40 times without any new primer or additional operation. This work provides new insight into understanding the reaction dynamic of dLAMP. The PIE strategy could be applied to the other digital amplification methods.