On-chip real-time single-copy polymerase chain reaction in picoliter droplets

Journal Article · Fri Apr 20 04:00:00 EDT 2007 · Analytical Chemistry, vol. 79, no. 22, November 15, 2007, pp. 8471-8475

DOI:https://doi.org/10.1021/ac701809w· OSTI ID:929191

Beer, N R; Hindson, B; Wheeler, E; Hall, S B; Rose, K A; Kennedy, I; Colston, B

The first lab-on-chip system for picoliter droplet generation and PCR amplification with real-time fluorescence detection has performed PCR in isolated droplets at volumes 10{sup 6} smaller than commercial real-time PCR systems. The system utilized a shearing T-junction in a silicon device to generate a stream of monodisperse picoliter droplets that were isolated from the microfluidic channel walls and each other by the oil phase carrier. An off-chip valving system stopped the droplets on-chip, allowing them to be thermal cycled through the PCR protocol without droplet motion. With this system a 10-pL droplet, encapsulating less than one copy of viral genomic DNA through Poisson statistics, showed real-time PCR amplification curves with a cycle threshold of {approx}18, twenty cycles earlier than commercial instruments. This combination of the established real-time PCR assay with digital microfluidics is ideal for isolating single-copy nucleic acids in a complex environment.

• Research Organization: Lawrence Livermore National Laboratory (LLNL), Livermore, CA
• Sponsoring Organization: USDOE
• DOE Contract Number: W-7405-ENG-48
• OSTI ID: 929191
• Report Number(s): UCRL-JRNL-230233
• Journal Information: Analytical Chemistry, vol. 79, no. 22, November 15, 2007, pp. 8471-8475, Journal Name: Analytical Chemistry, vol. 79, no. 22, November 15, 2007, pp. 8471-8475 Journal Issue: 22 Vol. 79
• Country of Publication: United States
• Language: English

Related Subjects

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY
47 OTHER INSTRUMENTATION
59 BASIC BIOLOGICAL SCIENCES
AMPLIFICATION
DETECTION
DNA
FLUORESCENCE
NUCLEIC ACIDS
POLYMERASE CHAIN REACTION
SILICON
STATISTICS