Microfluidics and Nanofluidics

, Volume 12, Issue 1–4, pp 423–429 | Cite as

Localized substance delivery to single cell and 4D imaging of its uptake using a flow channel with a lateral aperture

  • Kyohei Terao
  • Atsuhito Okonogi
  • Ariko Fuke
  • Teru Okitsu
  • Takaaki Suzuki
  • Masao Washizu
  • Hidetoshi Kotera
Research Paper

Abstract

We have developed a novel microfluidic device for space- and time-resolved (4D) visualization of intracellular events when a cell surface is partially exposed to external stimuli. The device, fabricated using 3D rotational inclined UV lithography of photoresist SU-8, consists of a cell-containing chamber and a flow channel separated by a thin vertical wall having a lateral micrometer aperture smaller than a cell. A cell is first immobilized on the aperture by suction from the flow channel using a syringe pump, and a chemical stimulant is then fed to the channel so that only the cell surface bounded by the aperture is subjected to the stimulus without leakage to other part of the cell surface. The subsequent lateral signal propagation inside the cell can be visualized using high-speed fluorescence confocal microscopy. As an experimental demonstration of the device, 2-NBDG (fluorescence glucose analog) intake into a mouse insulinoma cell, MIN6m9, was visualized in 4D resolution.

Keywords

Single cell analysis 4D imaging Microfluidic channel Localized substance delivery Multidirectional UV lithography 

Notes

Acknowledgments

We thank Prof. S. Seino for providing MIN6m9 cells. This work was supported in part by the research project ‘Development of Bio/Nano Hybrid Platform Technology towards Regenerative Medicine’, CREST, Japan Science and Technology Agency (JST). Part of this work was conducted in the Kyoto-Advanced Nanotechnology Network, supported by the ‘Nanotechnology Network’ of the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan.

References

  1. Bruus H (2008) Theoretical microfluidics. Oxford University Press, OxfordGoogle Scholar
  2. Dehmelt L, Bastiaens PIH (2010) Spatial organization of intracellular communication: insights from imaging. Nat Rev Mol Cell Biol 11(6):440–452CrossRefGoogle Scholar
  3. Eriksson E, Enger J, Nordlander B, Erjavec N, Ramser K, Goksor M, Hohmann S, Nystrom T, Hanstorp D (2006) A microfluidic system in combination with optical tweezers for analyzing rapid and reversible cytological alterations in single cells upon environmental changes. Lab Chip 7:71–76CrossRefGoogle Scholar
  4. Hamill OP, Martinac B (2001) Molecular basis of mechanotransduction in living cells. Physiol Rev 81(2):685–740Google Scholar
  5. Hammond AT, Glick BS (2000) Raising the speed limits for 4D fluorescence microscopy. Traffic 1(12):935–940CrossRefGoogle Scholar
  6. Han M, Lee W, Lee SK, Lee SS (2004) 3D microfabrication with inclined/rotated UV lithography. Sens Actuators A 111(1):14–20CrossRefGoogle Scholar
  7. Huang H, Kamm RD, Lee RT (2004) Cell mechanics and mechanotransduction: pathways, probes and phyiology. Am J Physiol Cell Physiol 287:C1–C11CrossRefGoogle Scholar
  8. Juncker D, Schmid H, Delamarche E (2005) Multipurpose microfluidic probe. Nat Mater 4(8):622–628CrossRefGoogle Scholar
  9. Khine M, Lau A, Ionescu-Zanetti C, Seo J, Lee LP (2005) A single cell electroporation chip. Lab Chip 5(1):38–43CrossRefGoogle Scholar
  10. Khine M, Ionescu-Zanetti C, Blatz A, Wang LP, Lee LP (2007) Single-cell electroporation arrays with real-time monitoring and feedback control. Lab Chip 7(4):457–462CrossRefGoogle Scholar
  11. Kuczenski B, Ruder WC, Messner WC, LeDuc PR (2009) Probing cellular dynamics with a chemical signal generator. PLoS One 4(3):0004847CrossRefGoogle Scholar
  12. Li CW, Yang J, Yang M (2006) Dose-dependent cell-based assays in V-shaped microfluidic channels. Lab Chip 6:921–929CrossRefGoogle Scholar
  13. Li X, Li PCH (2005) Microfluidic selection and retention of a single cardiac myocyte, on-chip dye loading, cell contraction by chemical stimulation, and quantitative fluorescent analysis of intracellular calcium. Anal Chem 77(14):4315–4322CrossRefGoogle Scholar
  14. Minami K, Yano H, Miki T, Nagashima K, Wang CZ, Tanaka H, Miyazaki JI, Seino S (2000) Insulin secretion and differential gene expression in glucose-responsive and -unresponsive MIN6 sublines. Am J Physiol Endocrinol Metabol 279(4):E773–E781Google Scholar
  15. Piper JD, Li C, Lo CJ, Berry R, Korchev Y, Ying LM, Klenerman D (2008) Characterization and application of controllable local chemical changes produced by reagent delivery from a nanopipet. J Am Chem Soc 130(31):10386–10393CrossRefGoogle Scholar
  16. Rorsman P, Renstrom E (2003) Insulin granule dynamics in pancreatic beta cells. Diabetologia 46(8):1029–1045CrossRefGoogle Scholar
  17. Routenberg DA, Reed MA (2009) Microfluidic probe: a new tool for integrating microfluidic environments and electronic wafer-probing. Lab Chip 10(1):123–127CrossRefGoogle Scholar
  18. Sato H, Yagyu D, Ito S, Shoji S (2006) Improved inclined multi-lithography using water as exposure medium and its 3D mixing microchannel application. Sens Actuators A 128(1):183–190CrossRefGoogle Scholar
  19. Sawano A, Takayama S, Matsuda M, Miyawaki A (2002) Lateral propagation of EGF signaling after local stimulation is dependent on receptor density. Dev Cell 3(2):245–257CrossRefGoogle Scholar
  20. Suzuki S, Yamamoto H, Ohoka M, Kanno I, Washizu M, Kotera H (2007) A low-damage cell trapping array fabricated by single-mask multidirectional photolithography with equivalent circuit analysis. Proc MicroTAS 2007(1):1765–1767Google Scholar
  21. Takayama S, Ostuni E, LeDuc P, Naruse K, Ingber DE, Whitesides GM (2001) Laminar flows—subcellular positioning of small molecules. Nature 411(6841):1016–1016Google Scholar
  22. Tavana H, Jovic A, Mosadegh B, Lee QY, Liu X, Luker KE, Luker GD, Weiss SJ, Takayama S (2009) Nanolitre liquid patterning in aqueous environments for spatially defined reagent delivery to mammalian cells. Nat Mater 8(9):736–741CrossRefGoogle Scholar
  23. Wheeler AR, Throndset WR, Whelan RJ, Leach AM, Zare RN, Liao YH, Farrell K, Manger ID, Daridon A (2003) Microfluidic device for single-cell analysis. Anal Chem 75(14):3581–3586CrossRefGoogle Scholar
  24. Yamada K, Nakata M, Horimoto N, Saito M, Matsuoka H, Inagaki N (2000) Measurement of glucose uptake and intracellular calcium concentration in single, living pancreatic beta-cells. J Biol Chem 275(29):22278–22283CrossRefGoogle Scholar
  25. Yoon YK, Park JH, Allen MG (2006) Multidirectional UV lithography for complex 3-D MEMS structures. J Microelectromech Syst 15(5):1121–1130CrossRefGoogle Scholar
  26. Zhu Z, Zhou ZF, Huang QA, Li WH (2008) Modeling, simulation and experimental verification of inclined UV lithography for SU-8 negative thick photoresists. J Micromech Microeng 18(12):125017CrossRefGoogle Scholar
  27. Zou C, Wang Y, Shen Z (2005) 2-NBDG as a fluorescent indicator for direct glucose uptake measurement. J Biochem Biophys Methods 64:207–215CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Kyohei Terao
    • 1
    • 2
  • Atsuhito Okonogi
    • 2
    • 3
  • Ariko Fuke
    • 3
  • Teru Okitsu
    • 2
    • 4
  • Takaaki Suzuki
    • 1
    • 2
  • Masao Washizu
    • 2
    • 5
  • Hidetoshi Kotera
    • 2
    • 3
  1. 1.Department of Intelligent Mechanical Systems EngineeringKagawa UniversityTakamatsuJapan
  2. 2.JST-CRESTSaitamaJapan
  3. 3.Department of Micro EngineeringKyoto UniversityKyotoJapan
  4. 4.Institute of Industrial ScienceThe University of TokyoTokyoJapan
  5. 5.Department of Mechanical EngineeringThe University of TokyoTokyoJapan

Personalised recommendations