JRM Vol.22 No.5 pp. 651-658
doi: 10.20965/jrm.2010.p0651


Quantitative Evaluation of Injected Molecules into Phospholipid-Coated Microdroplets for In situ Biological Reactions

Masahiro Nakajima*, Yuta Matsuno**, Masaru Kojima**,
Yohko Takiguchi***, Kingo Takiguchi***, Kousuke Nogawa**,
Michio Homma***, and Toshio Fukuda*,**

*Center For Micro-nano Mechatronics, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8603, Japan

**Department of Micro-Nano Systems Engineering, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8603, Japan

***Division of Biological Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya 464-8602, Japan

May 17, 2010
July 28, 2010
October 20, 2010
artificial cell model, micro/nanopipette, micromanipulation, phospholipid-coated microdroplet, biological reaction

This paper presents a quantitative evaluation of the amount of biological molecules injected into phospholipid-coated microdroplets. Research on developing an artificial cell model using lipid membrane vesicles has been pursued to determine the function between biomembranes and biological molecules. The method is needed to introduce biological molecules into the biomembrane model and observe their reactions. Conventionally, molecules are introduced into vesicles by hydrating dried lipid films or freeze-dried lipid blocks with a solution already including biological molecules. It is difficult, however, to observe reaction in real time and step-by-step for different types of biological molecules, because the reaction has already started by the time vesicles are formed. Our proposal uses micro/nanopipettes based on micromanipulation. It is demonstrated that the injection of different types of biological molecules into a phospholipid-coated microdroplet. Biological molecules, such as F-actin, heavy meromyosin (HMM), and adenosine triphosphate (ATP), were introduced into a phospholipidcoated microdroplet in sequence, and these reactions were observed inside the microdroplet. The amount of molecules introduced into the microdroplet was evaluated quantitatively from the intensity of fluorescent labels through our microchannel calibration method.

Cite this article as:
M. Nakajima, Y. Matsuno, M. Kojima, <. Takiguchi, K. Takiguchi, K. Nogawa, <. Homma, and T. Fukuda, “Quantitative Evaluation of Injected Molecules into Phospholipid-Coated Microdroplets for In situ Biological Reactions,” J. Robot. Mechatron., Vol.22, No.5, pp. 651-658, 2010.
Data files:
  1. [1] M. Hase, A. Yamada, T. Hamada, D. Baigl, and K. Yoshikawa, “Manipulation of Cell-Sized Phospholipid-Coated Microdroplets and Their Use as Biochemical Microreactors,” Langmuir, Vol.23, pp. 348-352, 2007.
  2. [2] M. Yamazaki and Y. Tamba, “The Single GUV Method for Probing Biomembrane Structure and Function,” J. of Surface Science and Nanotechnology, Vol. 3, pp. 218-227, 2005.
  3. [3] Y. Tamba, S. Ohba, M. Kubota, H. Yoshioka, and M. Yamazaki, “Single GUV Method Reveals Interaction of Tea Catechin (2)-Epigallocatechin Gallate with Lipid Membranes,” Biophysical J., Vol.92, pp. 3178-3194, 2007.
  4. [4] K. Takiguchi, A. Yamada, M. Negishi, Y. Tanaka-Takiguchi, and K. Yoshikawa, “Entrapping Desired Amounts of Actin Filaments and Molecular Motor Proteins in Giant Liposomes,” Langmuir, Vol.24, pp. 11323-11326, 2008.
  5. [5] M. Yamazaki, “The Single GUV Method to Reveal Elementaly Processes of Leakage of Internal Contents from Liposomes Induced by Antimicrobial Substances,” Adv. In Planar Lipiid Bilayers and Lipsomes, Vol.7, pp. 121-142, 2008.
  6. [6] Y. Tanaka-Takiguchi, M. Kinoshita, and K. Takiguchi, “Septinmediated Uniform Bracing of Phospholipid Membranes,” Current Biology, Vol.19, pp. 140-145, 2009.
  7. [7] M. Hase and K. Yoshikawa, “Structual Trasition of Actin Filament in a Cell-sized Water Droplet with a Phospholipid Membrane,” J. Chem. Phys., Vol.124, p. 104903, 2006.
  8. [8] M. Honda, K. Takiguchi, S. Ishikawa, and H. Hotani, “Morphogenesis of Liposomes Encapsulating Actin Depends on the Type of Actin-crosslinking,” J. of Molecular Biology, Vol.287, pp. 293-300, 1999.
  9. [9] D. A. Dean, “Import of Plasmid DNA into the Nucleus Is Sequence Specific,” Experimental Cell Research, Vol.230, pp. 293-302, 1997.
  10. [10] T. V. Tsulaia, N. L. Prokopishyn, A. Yao, N. D. Victor Carsrud, M. C. Carou, D. B. Brown, B. R. Davis, and J. Yannariello-Brown, “Glass Needle-Mediated Microinjection of Macromolecules and Transgenes into Primary Human Mesenchymal Stem Cells,” J. of biological science, Vol.10, pp. 328-336, 2003.
  11. [11] K. Nogawa, M. Kojima, M. Nakajima, S. Kojima, M. Homma, and T. Fukuda, “Rotational Speed Control of Na+-Driven Flagellar Motor by Dual Pipettes,” IEEE Trans. on Nanobiosicience, Vol.8, pp. 341-348, 2009.
  12. [12] Y. Matsuno, M. Nakajima, M. Kojima, Y. Tanaka-Takiguchi, K. Takiguchi, K. Nogawa, M. Homma, and T. Fukuda, “Pico-liter Injection Control to Individual Nano-liter Solution Coated by Lipid Layer,” Proc. of Int. Symp. on Micro-Nano Mechatronics and Human Science 2009, pp. 249-254, 2009.
  13. [13] M. Hase, A. Yamada, T. Hamada, D. Baigl, and K. Yoshikawa, “Manipulation of Cell-Sized Phospholipid-Coated Microdroplets and Their Use as Biochemical Microreactors,” Langmuir, Vol.23, pp. 348-352, 2007.
  14. [14] M. Nakajima, Y. Matsuno, Y. Tanaka-Takiguchi, K. Nogawa, K. Takiguchi, M. Homma, and T. Fukuda, “Evalution of Ultra-Minimal Spout Amount from Micro-Nano Pipettes into Phospholipid-Coated Micro-Droplet,” Trans. of JSME, Vol.76, No.766, pp. 1547-1552, 2010. (in Japanese)
  15. [15] I. Rayment, H. M. Holden, and M. Whittaker, “Structure of the Actin-Myosin Complex and Its Implications for Muscle Contraction,” Science, Vol.261, pp. 58-65, 1993.
  16. [16] S. Iwaia and T. Q. P. Uyeda, “Visualizing myosin-actin interaction with a genetically-encoded fluorescent strain sensor,” Proc. of the National Academy of Sciences, Vol.105, pp. 16882-16887, 2008.
  17. [17] A. Ishijima, H. Kojima, T. Funatsu, M. Tokunaga, H. Higuchi, H. Tanaka, and T. Yanagida, “Simultaneous Observation of Individual ATPase and Mechanical Events by a Single Myosin Molecule during Interaction with Actin,” Cell, Vol.92, pp. 161-171, 1998.

*This site is desgined based on HTML5 and CSS3 for modern browsers, e.g. Chrome, Firefox, Safari, Edge, Opera.

Last updated on Mar. 31, 2023