Site-specific immobilization of microbes using carbon nanotubes and dielectrophoretic force for microfluidic applications† Intae Kim, a Taechang An, b WooSeok Choi, a Chang Sup Kim, c Hyung Joon Cha c and Geunbae Lim * ad We developed a microbial immobilization method for successful applications in microfluidic devices. Single-walled nanotubes and Escherichia coli were aligned between two cantilever electrodes by a positive dielectrophoretic force resulting in a film of single-walled nanotubes with attached Escherichia coli. Because this film has a suspended and porous structure, it has a larger reaction area and higher reactant transfer efficiency than film attached to the substrate surface. The cell density of film was easily controlled by varying the cell concentration of the suspension and varying the electric field. The film showed excellent stability of enzyme activity, as demonstrated by measuring continuous reaction and long-term storage times using recombinant Escherichia coli that expressed organophosphorus hydrolase. Humans have used microorganisms before they even knew of their existence for such things as brewing and food fermentation. More recently, microbes that have special catalytic activity have been discovered and advancements in metabolic engineering and synthetic biology have enabled the use of microbes for specic metabolic processes and to synthesize specic enzymes. 1–5 Microbes have been successfully used for various applications, such as wastewater and soil treatment, biofuel cells, biofuel production, hydrogen production, and biosensors. 1,5–12 The use of microbes in microuidic applications has the potential to improve established technology and offer new methodologies in many elds. This is because microbes can be used in place of common enzymes in microuidic chips. 13,14 Microbes are a major host for enzyme production. Useable enzymes are produced as a result of a purication process because the enzymes are expressed in the intracellular space as an inclusion body. Recently, the secretion of enzymes into the periplasmic space or cell surface has been developed. 15,16 Using this technique, the enzymes in the periplasmic space or cell surface can come into contact with reactants without the need for a purication process. This means microbes can be used as whole-cell catalysts and can replace the use of enzymes. 17–21 In addition, the metabolic activity of microbes can be used in microuidic devices. Therefore, macroscale systems that use the metabolic activity of microbes can be miniaturized in microuidic devices and integrated in microuidic systems as bioreactors. To use microbes in microuidic applications, a microbial immobilization method that allows efficient xation and exact positioning in a small space is required. Various microbial cell immobilization methods, such as occulation, chemical attach- ment, gel entrapment, encapsulation, and biolms have been developed and are currently used. 7,12,22–24 However, these methods are unsuited for microuidic systems due to critical problems, such as mass transport limitations, difficulty in size reduction, non-site-specic immobilization, and slow processing. The present paper presents a one-step microbial immobili- zation method for microuidic applications using single-walled nanotubes (SWNTs), and dielectrophoretic (DEP) force. DEP force is an effective tool when manipulating biological cells and SWNTs. 25–28 This method is an extension of our lm fabrication technique involving dielectrophoretically aligned SWNTs. 28 In this method, SWNTs were functionalized by oxidation and mixed with an Escherichia coli (E. coli) suspension. The mixed suspension of SWNTs and E. coli cells was placed between two cantilever electrodes and the E. coli and SWNTs were aligned by the DEP force. Finally, an SWNT lm with attached E. coli was formed in the desired location. The effectiveness of this immobilization method was demonstrated by density control of the cells and the stability of cell activity. a Department of Mechanical Engineering, Pohang University of Science and Technology (POSTECH), San 31, Pohang, Gyungbuk, 790-784, The Republic of Korea. E-mail: limmems@postech.ac.kr b Department of Mechanical Design Engineering, Andong National University, Andong, Gyungbuk, 760-749, The Republic of Korea c Department of Chemical Engineering, Pohang University of Science and Technology (POSTECH), San 31, Pohang, Gyungbuk, 790-784, The Republic of Korea d Department of Integrative Bioscience and Biotechnology, Pohang University of Science and Technology (POSTECH), San 31, Pohang, Gyungbuk, 790-784, The Republic of Korea † Electronic supplementary information (ESI) available. See DOI: 10.1039/c3ra45155k Cite this: RSC Adv., 2014, 4, 1347 Received 16th September 2013 Accepted 18th November 2013 DOI: 10.1039/c3ra45155k www.rsc.org/advances This journal is © The Royal Society of Chemistry 2014 RSC Adv., 2014, 4, 1347–1351 | 1347 RSC Advances COMMUNICATION Published on 20 November 2013. Downloaded by Pohang University of Science and Technology on 02/06/2015 06:28:44. View Article Online View Journal | View Issue