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Summary
Digital PCR (dPCR) is a method used to quantify nucleic acids (DNA, RNA, cDNA). At NIST we are using microfluidic and emulsion-based dPCR platforms for quantification of viral and human genome SRMs.
Description
Digital PCR works through the partitioning of DNA molecules in an oil-emulsion droplet or physical chamber. The resulting PCR reactions will either be positive (presence of target) or negative (absence of target). By counting the positive and negative droplets, an estimate of the starting amount of DNA or RNA can be measured.
With appropriately validated assays, digital polymerase chain reaction (dPCR) determines the number of DNA targets (copies) per reaction partition, without the need for a standard curve. Reaction partitions can be in the form of fixed chambers in a microfluidic device, referred to as chamber digital PCR (cdPCR), or aqueous-based droplets floating in oil, referred to as droplet digital PCR (ddPCR). Once the chamber volume or the droplet volume is known the concentration of the measured sample can be determined as copies/µL.
In the Applied Genetics Group, the dPCR method is well-suited to the quantification of DNA and RNA-based reference materials.