Development of a microfluidics biosensor for agarose-bead immobilized Escherichia coli bioreporter cells for arsenite detection in aqueous samples

Abstract

Contamination with arsenic is a recurring problem in both industrialized and developing countries. Drinking water supplies for large populations can have concentrations much higher than the permissible levels (for most European countries and the United States, 10 μg As per L; elsewhere, 50 μg As per L). Arsenic analysis requires high-end instruments, which are largely unavailable in developing countries. Bioassays based on genetically engineered bacteria have been proposed as suitable alternatives but such tests would profit from better standardization and direct incorporation into sensing devices. The goal of this work was to develop and test microfluidic devices in which bacterial bioreporters could be embedded, exposed and reporter signals detected, as a further step towards a complete miniaturized bacterial biosensor. The signal element in the biosensor is a nonpathogenic laboratory strain of Escherichia coli, which produces a variant of the green fluorescent protein after contact to arsenite and arsenate. E. coli bioreporter cells were encapsulated in agarose beads and incorporated into a microfluidic device where they were captured in 500 × 500 μm(2) cages and exposed to aqueous samples containing arsenic. Cell-beads frozen at -20 °C in the microfluidic chip retained inducibility for up to a month and arsenic samples with 10 or 50 μg L(-1) could be reproducibly discriminated from the blank. In the 0-50 μg L(-1) range and with an exposure time of 200 minutes, the rate of signal increase was linearly proportional to the arsenic concentration. The time needed to reliably and reproducibly detect a concentration of 50 μg L(-1) was 75-120 minutes, and 120-180 minutes for a concentration of 10 μg L(-1).

Theory, fabrication and applications of microfluidic and nanofluidic biosensors

“…The substrate can then be patterned using other lithographic methods to produce more complicated structures [117]. Of these methods, nanoimprint lithography has been used by many researchers for the fabrication of PDMS fluidic channels for biosensors [3,11,25,26,45,61].…”

“…Common optical methods include fluorescence [3][4][5][6], optical cavity resonators [7][8][9] and surface-plasmon resonance or SPR [10][11][12][13][14][15][16] in which a surface-based chemical reaction corresponds to a change in the refractive index and thus a shift in the optical signal. Electrochemical impedance spectroscopy or EIS [17][18][19] detects changes in surface or nanostructure impedance owing to surface reactions or transport.…”

“…For example, a polymer-based disposable device capable of detecting 0.25-10 ppm of anaesthetic propofol within 60 s has been already demonstrated (figure 5) [114]. Poly(dimethyl siloxane) (PDMS) is the most commonly used polymer for fabrication of fluidic channels [3,4,11,60] towards biosensor applications though poly(methyl methacrylate) (PMMA) is also used [115]. Conducting polymers such as poly(phenylene vinylene) [23], and its soluble derivatives along with polyaniline have been used for electronic components [28].…”

“…Other target molecules include micro-RNA (miRNA) [24,28], a biomarker for cancer, DNA [15,20,25,30,34,61,122], herpes simplex-2 virus infection [5] and common drinking water contaminants including E. coli [45], pesticides [22,35], synthetic oestrogen in river water [21] and heavy metals [3,[162][163][164].…”

“…The solvents are usually aqueous solutions to preserve the natural state, or after processing may be organic materials. The sensor should be able to perform the [35][36][37] nanowires change in surface charge through protonation/deprotonation [33] electrical field-effect transistors changes in inherent biomolecular charge [38,[41][42][43]51] nanowires changes in electrical signal such as current, resistance, capacitance or conductance [24][25][26][27][28][29][30][31][32][33][34] optical fluorescence intensity of fluorescent signal [3][4][5] optical cavity resonator shift in resonance wavelength proportional to change in mass [7,9] surface-plasmon resonance shift in refractive index [10,11,[13][14][15][16]52] (b) sample manipulation method underlying principle involved references pumps electrokinetic pumping (including electro-osmosis) valves surface modification [53] droplets transfer electrically controlled surface tension drives liquid droplets [54,55] arrays/mixing of fluids use of heterogeneous surfaces (e.g. use of nanoholes or modified surfaces) [56,57] a As discussed in the text, many of the mechanisms can be combined with sample manipulation techniques in table 1b in microfluidic or nanofluidic platforms or can be used as stand-alone methods.…”