High-power white LED-based system incorporating a CCD Offner imaging spectrometer for real-time fluorescence qPCR measurements

Abstract

An optical system for qPCR fluorescence measurements which incorporates high-power white LEDs, PMMA plastic lenses and an Offner multichannel (imaging) CCD-based spectrometer has been developed and validated. The optical system can detect twenty reaction vessels in an asynchronous manner and up to seven different fluorescent dyes (7 plex) at 1 nM dye concentrations in each of the reaction vessels. Furthermore, PCR curves obtained using the optical measurement system for a genomic deoxyribonucleic acid (DNA) template containing HEX and Texas Red fluorescent probes (fluorophores) are discussed. The spectral resolution, dynamic range and repeatability of the measurement system are < 15 nm, > 3 decades and < 1% CV, respectively.