June 1, 2022
In this webinar Dr. Jan Hellemans, CTO and co-founder of Biogazelle, a CellCarta company, describes the intrinsic potential of droplet digital PCR (ddPCR) in clinical research for three different applications:
- rare variant analysis in plasma samples,
- small changes in alternative splicing, and
- CAR-T studies.
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Since becoming commercially available in 2011, digital PCR (dPCR) has been gaining in popularity as it enables nucleic acid quantification with superior accuracy and precision compared to quantitative PCR (qPCR). In brief, the technology consists of the following steps:
- The PCR reaction mixture is partitioned into thousands of water-in-oil droplets with target and background DNA randomly distributed among the reactions.
- The target DNA is amplified by PCR using standard thermal cycling with fluorescent dyes or probes.
- Each reaction provides a fluorescent positive or negative signal indicating that target DNA was present or absent in partitioning.
- The fraction of positive droplets is used to calculate the target DNA concentration using Poisson correction.
With data acquisition taking place at the end of the reaction, and since absolute concentrations can be determined, data can be easily interpreted and compared across different samples and experiments, especially in a clinical setting. Additionally, dPCR doesn’t suffer from the inherent complications of qPCR that result from the requirement of a standard curve and excels in the quantification of low abundant genes or small differences.