title: Novel fiber optic detection method for in situ analysis of fluorescently labeled biosensor organisms.
abstract: This research presents a novel, time-resolved fiber-optic "Optrode" system for accurate real-time in situ detection of fluorescent proteins produced by biosensor organisms. The Optrode fluorescence detection system was able to identify, characterize, differentiate, and quantify red and green fluorescently labeled organisms, individually and in mixed aqueous cultures. Detection was also possible in sand systems, where a consistent reduction in signal intensity indicates that signal collection volume was reduced by one-third. The optrode was shown to be sensitive enough to detect fluorescently labeled cell concentrations of 1.9 × 10(4) CFU/mL, indicating it is suitable for detecting typical concentrations of degrader organisms reported in bioremediation trials. The effect of fluorophore photobleaching was characterized for different fluorescent proteins and demonstrated a linear relationship to cell concentration, meaning the effect can be accounted for within methods of fluorescence collection and analysis. Proof of concept is provided for all aspects of this research, which represents an important step toward the goal of achieving a complete, nondestructive, in situ monitoring system to characterize all aspects of microbial activity and gene expression.
authors.affiliation: Department of Civil and Environmental Engineering, University of Auckland, New Zealand.
authors.forename: Belinda M
authors.initials: BM
authors.lastname: Hewitt
authors.forename: Naresh
authors.initials: N
authors.lastname: Singhal
authors.forename: Roy G
authors.initials: RG
authors.lastname: Elliot
authors.forename: Andy Y H
authors.initials: AY
authors.lastname: Chen
authors.forename: Jason Y C
authors.initials: JY
authors.lastname: Kuo
authors.forename: Frédérique
authors.initials: F
authors.lastname: Vanholsbeeck
authors.forename: Simon
authors.initials: S
authors.lastname: Swift
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D004352:Drug Resistance, Microbial; D004926:Escherichia coli / Q000378:metabolism*; D005336:Fiber Optic Technology / Q000379:methods*; D017930:Genes, Reporter / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000378:metabolism; D057230:Limit of Detection; D008164:Luminescent Proteins / Q000378:metabolism; D055100:Optical Fibers; D038761:Photobleaching; D016958:Pseudomonas putida / Q000302:isolation & purification / Q000378:metabolism*; D059629:Signal-To-Noise Ratio; D012965:Sodium Chloride; D012988:Soil Microbiology; D013050:Spectrometry, Fluorescence; D013194:Staining and Labeling
chemical_list: D008164:Luminescent Proteins; C413663:fluorescent protein 583; D049452:Green Fluorescent Proteins; D012965:Sodium Chloride