Digital PCR for higher accuracy, sensitivity and absolute quantification

Digital PCR is a new approach to nucleic acid detection and quantification that offers an alternate method to conventional real-time quantitative PCR for absolute quantification and rare allele detection. Digital PCR works by partitioning a sample of DNA or cDNA into many individual, parallel PCR reactions; some of these reactions contain the target molecule (positive) while others do not (negative). A single molecule can be amplified a million-fold or more. During amplification, TaqMan chemistry with dye-labeled probes is used to detect sequence-specific targets. When no target sequence is present, no signal accumulates. Following PCR analysis, the fraction of negative reactions is used to generate an absolute count of the number of target molecules in the sample, without the need for standards or endogenous controls.

The use of a nanofluidic chip provides a convenient and straightforward mechanism to run thousands of PCR reactions in parallel. Each well is loaded with a mixture of sample, master mix, and Applied Biosystems TaqMan Assay reagents, and individually analyzed to detect the presence (positive) or absence (negative) of an endpoint signal. To account for wells that may have received more than one molecule of the target sequence, a correction factor is applied using the Poisson model.  

 

Digital PCR
Advantages of digital PCR
  • No need to rely on references or standards
  • Ability to increase precision by using more PCR replicates
  • High tolerance to inhibitors
  • Capability to analyze complex mixtures
  • Linear detection of small-fold changes

Compare digital PCR and real-time PCR ›

Digital PCR extends capabilities for advanced applications

Digital PCR instruments and reagents enhance the performance of existing TaqMan Assays, enabling applications that benefit from higher sensitivity, precision, and absolute quantification. 

Absolute quantification, using sealed-chip technology, for a reliable method and precise, sensitive data.

Find the reagents and consumables you need for absolute quantification using digital PCR.

Digital PCR applications

Detect and quantify small percent copy number differences with a high degree of precision.

Detect and quantify rare mutations for low-prevalence targets in cancer research samples.

Absolute quantification of bacterial and viral loads, and absolute pathogen counts.

Generate absolute reference standards for genetic measurements, metrology, and cross-lab comparisons.

Absolute quantification of NGS libraries and validation of sequencing results, without reference standards.

Detect gene expressions changes of ±10% for absolute transcript quantification without a reference gene.

Detect low-level pathogens that cause human illnesses through contaminated food and water supplies.

Enables sensitive detection and absolute quantification of plant mutations and genetically modified organisms.

Applied Biosystems QuantStudio 3D Digital PCR System

  • Absolute data with high precision and sensitivity—detect and count individual molecules
  • Simple workflow—just load and go for digital data from chip-based system
  • Affordable platform—start doing digital PCR immediately, and at an affordable cost
  • Compatible—use your existing TaqMan real-time PCR assays
  • Sealed system—helps limit contaminants using a sealed chip without exposed sample-transfer steps
  • Wide dynamic range—achieve linearity over 5 logs
  • Intuitive—view data directly on touch screen and analyze efficiently using Applied Biosystems AnalysisSuite Digital PCR Software 

 

Resources

 

Support

 

For Research Use Only. Not for use in diagnostic procedures.