Droplet digital PCR of viral ‎DNA/RNA, current progress, challenges, and future perspectives

Kojabad, Amir Asri; Farzanehpour, Mahdieh; Galeh, Hadi Esmaeili Gouvarchin; Dorostkar, Ruhollah; Jafarpour, Ali; Bolandian, Masoumeh; Nodooshan, Majid Mirzaei

High‐throughput droplet‐based digital PCR (ddPCR) is a refinement of the conventional polymerase chain reaction (PCR)‎ methods. In ddPCR, DNA/RNA is encapsulated stochastically inside the microdroplets as reaction chambers. A small percentage of the reaction chamber contains one or fewer copies of the DNA or RNA. After PCR amplification, concentrations are determined based on the proportion of nonfluorescent partitions through the Poisson distribution. Some of the main features of ddPCR include high sensitivity and specificity, absolute quantification without a standard curve, high reproducibility, good tolerance to PCR inhibitor, and high efficacy compared to conventional molecular methods. These advantages make ddPCR a valuable addition to the virologist's toolbox. The following review outlines the recent technological advances in ddPCR methods and their applications in viral identification.