Mr. Jeffrey A. Goldman Profile

Jeffrey A. Goldman

Scientist at Bio-Rad Labs Inc

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Proceedings Article | 21 June 2002 Paper

Continuous fluorescence detection thermal cycling for DNA analysis using an array of LED light sources

Jeffrey Goldman, Daniel Sullivan, Igor Kordunskiy

Proceedings Volume 4626, (2002) https://doi.org/10.1117/12.472088

KEYWORDS: Light emitting diodes, Luminescence, Fluorescence spectroscopy, Light sources, Molecules, Calibration, Signal detection, Control systems, Photomultipliers, Imaging systems

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Numerous commercial instruments exist for assaying fluorescence signals from an array of samples during thermal cycling and thermal denaturation profiling; however these instruments tend to be complex, expensive, and/or require custom reaction vessels. We have developed a system that incorporates a novel fluorescence excitation scheme in which an independently addressable array of light sources is imaged into the wells of a standard 96-well plate in a sequential fashion and the resultant fluorescent excitation is read by a photomultiplier tube (PMT). By individually illuminating each sample, we have eliminated the need for sophisticated detection and image analysis techniques used to discriminate signals form different samples. Additionally, independent control of each light source allows the system to automatically balance the light incident on each vessel to provide a flat response across the array, thus avoiding any post-measurement normalization that would reduce dynamic range. The resulting instrument has high sensitivity due to the PMT, but low sample crosstalk due to the sample illumination control granted by the LED array. Application data is shown including single copy DNA amplification and detection as well as product thermal denaturation analysis.

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