Kanagawa Accademy of Science and Technology
Artificial Cell Membrane Systems Group

Brief summary

Biological membranes contain a wide variety of membrane proteins that carry out important physiological processes such as the transport of essential nutrients, signal transduction and neurotransmission.
Project Leader: Dr. Shoji Takeuchi (Assoc. prof. of Univ. of Tokyo)
Investigation period: 2009-2013.
Membrane proteins are the target of over half of all pharmaceutical drugs. Despite the importance of membrane proteins, researchers must address the formidable problem that, in most cases, these proteins preserve their original structure and activity within the membrane. Our project team has therefore been studying methods for reconstituting membrane proteins in artificial lipid bilayer membrane systems. In particular, we have focused on launching high throughput screening platforms for ion channels, receptors and transporters that would help expedite and decrease the cost of drug discovery. We utilize MEMS (Micro Electro Mechanical Systems) technologies to fabricate microchips that reproduce cellular membranes in vitro.

Project details

1. Planar bilayer lipid membrane systems for ion channel signal recordings

The droplet contact method (DCM), which we were the first to report, is one of the most simple, robust and reproducible methods for forming a planar bilayer lipid membrane (BLM) [Funakoshi, et al. Anal. Chem. 2006]. The DCM provides a planar BLM at the interface between two aqueous droplets in organic solvent that contains a lipid suspension and makes use of the self-assembling process of amphiphilic lipid molecules at the surface of the two aqueous droplets (Fig. 1). Since the method provides a BLM with a sufficiently high electrical resistance, the device is suitable for signal recording of a single ion channel protein at the picoampere level. Furthermore, the simple formation procedure (i.e., dropping aqueous solutions and a lipid-dispersed solvent sequentially) is amenable to automation and mass production of BLMs on a chip. We have published several reports demonstrating the principle of the DCM (Osaki, et al. Anal. Chem. 2009, Kawano, et al. Small 2010, Kawano, et al. J. Am. Chem. Soc. 2011).

Fig. 1 A conceptual diagram of the droplet contact method (DCM) and the BLM formation steps (right images).


2. Liposomal membrane systems for fluorescence monitoring of transporters and receptors

Fluorescence microscope observations of liposomal membrane platforms have shed light on molecular transport through membrane transporter/ligand binding to a membrane receptor. Membrane transport phenomena are difficult to monitor with electrical recordings. The formation of uniform-sized liposomes is a major challenge in the development of liposomal membrane platforms, but size uniformity is necessary for statistical analyses. We thus introduced a method that produces cell-sized liposomes in a microfluidic channel [Ota, et al. Angew. Chem. 2009]. This method, called blowing liposome formation, applies a fluid (jet) flow to a planar BLM that is formed using the DCM (Fig. 2). Another problem in studying membrane transport is the need to immobilize the liposomes. We recently demonstrated the advantage of a DNA anchoring technique to immobilize proteoliposomes onto a microfluidic channel, allowing a quantitative and statistical assay of a transporter protein, MDR1, under a fluorescence microscope [Sasaki, et al. Lab Chip 2012].

Fig. 2 Fluorescence microscope images of blowing liposome formation in a microfluidic channel. A planar BLM was pushed out from bottom to top, and a liposome was formed by shear stress in the flow channel (left to right). Formed liposomes are also shown. The inner solution contains a fluorescent molecule, calcein.