Effect of substrate thermal resistance on space-domain microchannel fluorescent melting curve analysis

Biomed Microdevices. 2009 Aug;11(4):747-54. doi: 10.1007/s10544-009-9288-6.

Abstract

In recent years, Fluorescent Melting Curve Analysis (FMCA) has become an almost ubiquitous feature of commercial quantitative PCR (qPCR) thermal cyclers. Here a micro-fluidic device is presented capable of performing FMCA within a microchannel. The device consists of modular thermally conductive blocks which can sandwich a microfluidic substrate. Opposing ends of the blocks are held at differing temperatures and a linear thermal gradient is generated along the microfluidic channel. Fluorescent measurements taken from a sample as it passes along the micro-fluidic channel permits fluorescent melting curves to be generated. In this study we measure DNA melting temperature from two plasmid fragments. The effects of flow velocity and ramp-rate are investigated, and measured melting curves are compared to those acquired from a commercially available PCR thermocycler.

MeSH terms

  • Fluorescence
  • Hot Temperature
  • Microfluidic Analytical Techniques / instrumentation*
  • Microfluidic Analytical Techniques / methods*
  • Plasmids / chemistry*
  • Polymerase Chain Reaction / instrumentation*
  • Polymerase Chain Reaction / methods*
  • Sequence Analysis, DNA / instrumentation
  • Sequence Analysis, DNA / methods