High-Throughput Droplet Digital PCR System for Absolute Quantitation of DNA Copy Number

2011/10/28 by Benjamin J. Hindson, Kevin D. Ness, Donald A Masquelier +42 · 2,817 citations
Biochemistry, Genetics and Molecular Biology · Chemistry · Medicine · #Biochemistry #Biology #Cancer Genomics and Diagnostics #Chemistry #Computational biology #Computer science #DNA #Digital polymerase chain reaction #Gene #Nucleic acid #Polymerase chain reaction #Prenatal Screening and Diagnostics #Real-time polymerase chain reaction #Single-cell and spatial transcriptomics #TaqMan #Throughput

paper · pdf · doi:10.1021/ac202028g

published in Analytical Chemistry 83(22), 8604-8610 (American Chemical Society)

openalex publication_date 2011/10/28 · openalex created_date 2025/10/10 · openalex updated_date 2026/08/05

Abstract

Digital PCR enables the absolute quantitation of nucleic acids in a sample. The lack of scalable and practical technologies for digital PCR implementation has hampered the widespread adoption of this inherently powerful technique. Here we describe a high-throughput droplet digital PCR (ddPCR) system that enables processing of ~2 million PCR reactions using conventional TaqMan assays with a 96-well plate workflow. Three applications demonstrate that the massive partitioning afforded by our ddPCR system provides orders of magnitude more precision and sensitivity than real-time PCR. First, we show the accurate measurement of germline copy number variation. Second, for rare alleles, we show sensitive detection of mutant DNA in a 100,000-fold excess of wildtype background. Third, we demonstrate absolute quantitation of circulating fetal and maternal DNA from cell-free plasma. We anticipate this ddPCR system will allow researchers to explore complex genetic landscapes, discover and validate new disease associations, and define a new era of molecular diagnostics.

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