Biofilm formation in microfluidic channels is difficult to detect because sampling volumes are too small for conventional turbidity measurements. To detect biofilm formation, we used an ion-sensitive field-effect transistor (ISFET) measurement system to measure pH changes in small volumes of bacterial suspension. Cells of Micrococcus luteus ( M. luteus) were cultured in polystyrene (PS) microtubes and polymethylmethacrylate (PMMA)-based microfluidic channels laminated with polyvinylidene chloride. In microtubes, concentrations of bacteria and pH in the suspension were analyzed by measuring turbidity and using an ISFET sensor, respectively. In microfluidic channels containing 20 μL of bacterial suspension, we measured pH changes using the ISFET sensor and monitored biofilm formation using a microscope. We detected acidification and alkalinization phases of M. luteus from the ISFET sensor signals in both microtubes and microfluidic channels. In the alkalinization phase, after 2 day culture, dense biofilm formation was observed at the bottom of the microfluidic channels. In this study, we used an ISFET sensor to detect biofilm formation in clinical and industrial microfluidic environments by detecting alkalinization of the culture medium.
References
[1]
Chávez de Paz, L.E.; Hamilton, I.R.; Svens？ter, G. Oral bacteria in biofilms exhibit slow reactivation from nutrient deprivation. Microbiology 2008, 154, 1927–1938.
[2]
Coenye, T.; De Prijck, K.; De Wever, B.; Nelis, H.J. Use of the modified Robbins device to study the in vitro biofilm removal efficacy of NitrAdine？, a novel disinfecting formula for the maintenance of oral medical devices. J. Appl. Microbiol. 2008, 105, 733–740.
[3]
Eighmy, T.T.; Maratea, D.; Bishop, P.L. Electron microscopic examination of wastewater biofilm formation and structural components. Appl. Environ. Microbiol. 1983, 45, 1921–1931.
[4]
Chmielewski, R.A.N.; Frank, J.F. Biofilm formation and control in food processing facilities. Comprehens. Rev. Food Sci. Food Saf. 2003, 2, 22–32.
[5]
Lee, J.-H.; Kaplan, J.B.; Lee, W.Y. Microfluidic devices for studying growth and detachment of Staphylococcus epidermidis biofilms. Biomed. Microdevices 2008, 10, 489–498.
[6]
Yawata, Y.; Toda, K.; Setoyama, E.; Fukuda, J.; Suzuki, H.; Uchiyama, H.; Nomura, N. Monitoring biofilm development in a microfluidic device using modified confocal reflection microscopy. J. Biosci. Bioeng. 2010, 110, 377–380.
[7]
Bos, R.; van der Mei, H.C.; Busscher, H.J. Physico-chemistry of initial microbial adhesive interactions—Its mechanisms and methods for study. FEMS Microbiol. Rev. 1999, 23, 179–230.
[8]
Ponsonnet, L.; Boureanu, M.; Jaffrezic, N.; Othmane, A.; Dorel, C.; Lejeune, P. Local pH variation as an initial step in bacterial surface-sensing and biofilm formation. Mater. Sci. Eng. C 2008, 28, 896–900.
[9]
Burne, R.A.; Marquis, R.E. Alkali production by oral bacteria and protection against dental caries. FEMS Microbiol. Lett. 2000, 193, 1–6.
[10]
Bergveld, P. Thirty years of ISFETOLOGY: What happened in the past 30 years and what may happen in the next 30 years. Sens. Actuators B Chem. 2003, 88, 1–20.
[11]
Lee, C.S.; Kim, S.K.; Kim, M. Ion-sensitive field-effect transistor for biological sensing. Sensors 2009, 9, 7111–7131.
[12]
Wang, J.; Wu, C.; Hu, N.; Zhou, J.; Du, L.; Wang, P. Microfabricated electrochemical cell-based biosensors for analysis of living cells in vitro. Biosensors 2012, 2, 127–170.
[13]
Mohri, S.; Shimizu, J.; Goda, N.; Miyasaka, T.; Fujita, A.; Nakamura, M.; Kajiya, F. Measurements of CO2, lactic acid and sodium bicarbonate secreted by cultured cells using a flow-through type pH/CO2 sensor system based on ISFET. Sens. Actuators B Chem. 2006, 115, 519–525.
[14]
Mohri, S.; Yamada, A.; Goda, N.; Nakamura, M.; Naruse, K.; Kajiya, F. Application of a flow-through type pH/CO2 sensor system based on ISFET for evaluation of the glucose dependency of the metabolic pathways in cultured cells. Sens. Actuators B Chem. 2008, 134, 447–450.
[15]
Sakata, T.; Makino, I.; Kita, S. Real-time and noninvasive monitoring of respiration activity of fertilized ova using semiconductor-based biosensing devices. Eur. Biophys. J. 2011, 40, 699–704.
[16]
Pourciel-Gouzy, M.L.; Assié-Souleille, S.; Mazenq, L.; Launay, J.; Temple-Boyer, P. pH-ChemFET-based analysis devices for the bacterial activity monitoring. Sens. Actuators B Chem. 2008, 134, 339–344.
[17]
Pourciel-Gouzy, M.L.; Sant, W.; Humenyuk, I.; Malaquin, L.; Dollat, X.; Temple-Boyer, P. Development of pH-ISFET sensors for the detection of bacterial activity. Sens. Actuators B Chem. 2004, 103, 247–251.
[18]
Mayanagi, G.; Igarashi, K.; Washio, J.; Nakajo, K.; Domon-Tawaraya, H.; Takahashi, N. Evaluation of pH at the bacteria-dental cement interface. J. Dent. Res. 2011, 90, 1446–1450.
[19]
Volle, C.B.; Ferguson, M.A.; Aidala, K.E.; Spain, E.M.; Nú？ez, M.E. Spring constants and adhesive properties of native bacterial biofilm cells measured by atomic force microscopy. Colloids. Surf. B Biointerf. 2008, 67, 32–40.
[20]
Matsuura, K.; Uozumi, T.; Furuichi, T.; Sugimoto, I.; Kodama, M.; Funahashi, H. A Microfluidic device to reduce treatment time of intracytoplasmic sperm injection. Fertil. Steril. 2013, 99, 400–407.
[21]
Das, P.; Mukherjee, S.; Sen, R. Antimicrobial potential of a lipopeptide biosurfactant derived from a marine Bacillus circulans. J. Appl. Microbiol. 2008, 104, 1675–1684.
[22]
Yamada, A.; Mohri, S.; Nakamura, M.; Naruse, K. A fully automated pH measurement system for 96-well microplates using a semiconductor-based pH sensor. Sens. Actuators B Chem. 2010, 143, 464–469.
[23]
Blankenhorn, D.; Phillips, J.; Slonczewski, J.L. Acid- and base-induced proteins during aerobic and anaerobic growth of Escherichia coli revealed by two-dimensional gel electrophoresis. J. Bacteriol. 1999, 181, 2209–2216.
[24]
Hu, M.; Zhang, C.; Mu, Y.; Shen, Q.; Feng, Y. Indole affects biofilm formation in bacteria. Indian J. Microbiol. 2010, 50, 362–368.
[25]
Weeks, D.L.; Eskandari, S.; Scott, D.R.; Sachs, G. A H+-gated urea channel: The link between Helicobacter pylori urease and gastric colonization. Science 2000, 287, 482–485.
[26]
Lazar Ultra-M Micro pH Electrode Recommended by Researchers Worldwide, Available online: http://www.shelfscientific.com/cgi-bin/tame/newlaz/microphnsys.tam/ (accessed on 1 December 2012).
