Development of a flow-through detector for monitoring genotoxic compounds by quantifying the expression of green fluorescent protein in genetically modified yeast cells

Abstract

A flow-through detector has been developed for quantifying green fluorescent protein (GFP) expressed in genetically modified yeast cells upon exposure to a genotoxic compound. The detector forms the basis of a prototype semi-automated bioassay for genotoxic compounds, by biological fluorescence induction and detection. The selection and optimization of the various optical components of the system are discussed. The complete system was successfully tested with a known DNA damaging agent, and was able to monitor the expression of GFP on-line, in situ and in real time.

Green fluorescent protein inSaccharomyces cerevisiae: Real-time studies of theGAL1 promoter

“…Other than E. coli, GFP has also been used to monitor protein production in insect cells (Cha et al, 1997). Meanwhile, various methods and devices for quantitative measurements of GFP were developed (Randers-Eichhorn et al, 1997;Craig et al, 1997;Korf et al, 1997;Endow and Piston, 1998;Knight et al, 1999).…”

All solid-state GFP sensor

“…Another problem is the selectivity of the sensors (Knight et al, 1999). Fermentation media are usually highly fluorescent, with very broad emission spectra.…”

“…Optical fibers strongly attenuate both the excitation light and the output fluorescence and photomultipliers are used for light detection (Craig et al, 1998;Knight et al, 1999;Korf et al, 1997;Randers-Eichhorn et al, 1997). This imposes the need for specific design that provides very good optical shielding in order to avoid damage to the photodetector.…”

Optical tracking of a stress-responsive gene amplifier applied to cell-based biosensing and the study of synthetic architectures

“…doi:10.1016/j.bios.2009.12.036 candidates for miniature or remotely deployable assays. Namely, high reporter stability, signal accumulation, absence of a requirement for accessory substrates, and the non-invasiveness of the optical probing enable remote or miniature sensing applications (Baumstark-Khan et al, 2001, 2003Belkin, 2003;Knight et al, 1999;Kostov et al, 2000;Kostrzynska et al, 2002;Kuang et al, 2004;Lichtenberg-Fraté et al, 2003;Norman et al, 2005;Rabbow et al, 2003;Sagi et al, 2003;Zanzotto et al, 2006).…”