title: Polyacrylamide hydrogel encapsulated E. coli expressing metal-sensing green fluorescent protein as a potential tool for copper ion determination.
abstract: A simple, inexpensive and field applicable metal determination system would be a powerful tool for the efficient control of metal ion contamination in various sources e.g. drinking-water, water reservoir and waste discharges. In this study, we developed a cell-based metal sensor for specific and real-time detection of copper ions. E. coli expressing metal-sensing green fluorescent protein (designated as TG1/(CG)6GFP and TG1/H6CdBP4GFP) were constructed and served as a metal analytical system. Copper ions were found to exert a fluorescence quenching effect, while zinc and cadmium ions caused minor fluorescence enhancement in the engineered bacterial suspension. To construct a user-friendly and reagentless metal detection system, TG1/H6CdBP4GFP and TG1/(CG)6GFP were encapsulated in polyacrylamide hydrogels that were subsequently immobilized on an optical fiber equipped with a fluorescence detection module. The sensor could be applied to measure metal ions by simply dipping the encapsulated bacteria into a metal solution and monitoring fluorescence changes in real time as a function of the metal concentration in solution. The sensor system demonstrated high specificity toward copper ions. The fluorescence intensities of the encapsulated TG1/(CG)6GFP and TG1/H6CdBP4GFP were quenched by approximately 70 % and 80 % by a high-dose of copper ions (50 mM), respectively. The level of fluorescence quenching exhibited a direct correlation with the copper concentration, with a linear correlation coefficient (r) of 0.99. The cell-based metal sensor was able to efficiently monitor copper concentrations ranging between 5 M and 50 mM, encompassing the maximum allowed copper contamination in drinking water (31.15 M) established by the WHO. Furthermore, the cell-based metal sensor could undergo prolonged storage for at least 2 weeks without significantly influencing the copper sensitivity.
authors.affiliation: Center for Innovation Development and Technology Transfer, Faculty of Medical Technology, Mahidol University, Bangkok 10700, Thailand.
authors.forename: Tanawut
authors.initials: T
authors.lastname: Tantimongcolwat
authors.affiliation: Department of Clinical Microbiology and Applied Technology, Faculty of Medical Technology, Mahidol University, Bangkok 10700, Thailand.
authors.forename: Chartchalerm
authors.initials: C
authors.lastname: Isarankura-Na-Ayudhya
authors.affiliation: Department of Clinical Microscopy, Faculty of Medical Technology, Mahidol University, Bangkok 10700, Thailand.
authors.forename: Apapan
authors.initials: A
authors.lastname: Srisarin
authors.affiliation: Institute of Biochemistry, University of Muenster, Wilhelm-Klemm-Str. 2, Muenster 48149, Germany.
authors.forename: Hans-Joachim
authors.initials: HJ
authors.lastname: Galla
authors.affiliation: Department of Clinical Microbiology and Applied Technology, Faculty of Medical Technology, Mahidol University, Bangkok 10700, Thailand.
authors.forename: Virapong
authors.initials: V
authors.lastname: Prachayasittikul
keywords: copper; green fluorescent protein; metal sensor; optical sensor