title: A perfusable microfluidic device with on-chip total internal reflection fluorescence microscopy (TIRFM) for in situ and real-time monitoring of live cells.
abstract: A microfluidic device integrated with a Total Internal Reflection (TIR)-based chip for cell observation and analysis was developed. This integrated device enables in situ Total Internal Reflection Fluorescence Microscopy (TIRFM) on adherent cells cultured under continuous medium perfusion. This TIR-based chip, allows TIRFM to be easily performed on cells without the assembly of complicated optical components and cell culture chambers. The integrated device was evaluated by tracking the movement of fluorescent beads and monitoring the location of insulin granules in mouse pancreatic β-cells. This system offers higher signal-to-noise (S/N) ratio than epi-fluorescence microscopy (EPIFM), and comparable image quality to commercial TIRFM systems when imaging insulin granules. We also detected repetitive changes in intracellular Ca(2+) concentration in MIN6-m9 cells stimulated with KCl, which demonstrates quick perfusion for cell analysis while maintaining high S/N ratio.
authors.affiliation: Department of Microengineering, Kyoto University, Sakyo-ku, Kyoto, Japan. ryuji@me.kyoto-u.ac.jp
authors.forename: Ryuji
authors.initials: R
authors.lastname: Yokokawa
authors.forename: Yuko
authors.initials: Y
authors.lastname: Kitazawa
authors.forename: Kyohei
authors.initials: K
authors.lastname: Terao
authors.forename: Atsuhito
authors.initials: A
authors.lastname: Okonogi
authors.forename: Isaku
authors.initials: I
authors.lastname: Kanno
authors.forename: Hidetoshi
authors.initials: H
authors.lastname: Kotera
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D007328:Insulin / Q000378:metabolism; D051379:Mice; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D008856:Microscopy, Fluorescence / Q000295:instrumentation*; D008863:Microspheres; D057054:Molecular Imaging / Q000295:instrumentation*; D010477:Perfusion / Q000295:instrumentation*; D013997:Time Factors
chemical_list: D007328:Insulin; D002118:Calcium