The dimensions of the systems have been drastically scaled down in the last decades. The increasing demand of this miniaturization has brought about several challenges such as handling and transport of tiny amounts of materials. Microfluidics, a popular method, is not always appropriate when the amount of molecules to be handled is extremely small. Direct transport, as in intracellular transport, has potential to overcome the handling challenges due to the scaling down technology.

In this work, a heterogeneous integrated system, i.e. bio/MEMS hybrid system, has been proposed to build a nano-scale transport system based on kinesin motion along immobilized microtubules. In such a system, specificity and throughput of the bio-world can be successfully integrated with the handling and manipulation capabilities of MEMS technology to deliver systems capable of working at the nano-scale with high efficiency.

Kinesin is a molecular motor providing unidirectional motion on rail structures called microtubules. Orientation of the rail structures provides a controlled transport system. Although there have been some research on the microtubule assembly, so far, no technique could provide enough freedom to build a complex multidirectional microtubule network. However, to build a real nano system with a high level of transport capabilities, multidirectional assembly in a very small area is essential.

This research proposes an innovative method to build a multidirectional and multilayered assembly of microtubule networks. The technique was named as Picking and Relocation of Individual Microtubule Process abbreviated as PRIM Process. This method was based on individual manipulation of polarity-marked microtubules using MEMS tweezers. Isolated single microtubules were individually captured and then relocated on a predefined docking zone. As a result, due to the high-degree of freedom provided by direct manipulation of microtubules, multidirectional orientation and stacking was achieved in a very small area. Furthermore, it is demonstrated that micrometer precision could be achieved to relocate the captured microtubules onto predefined areas. Even three-dimensional relocation was performed to combine microtubule networks in different layers on a chip. Kinesin-coated bead motion on top of the assembled microtubules was successfully realized proving that the assembly method did not disturb the kinesin activity.

Building a system to handle target molecules requires a capturing process. Biofunctional beads were used to realize this process. Based on specific interactions between the target molecules and biofunctional coating of the beads, very efficient capturing could be achieved. To avoid any possible hindrance of kinesin motion and good integration with the PRIM Process, the transport mechanism was separated from the capturing mechanism. Several different carriers, such as beads, oil droplets, lipid vesicles, needles were adopted and successful transport was shown. To show the efficiency of the sorting and transport mechanism, a sorting device was built. Two different target molecules (biotin & antibody) were captured separately on two kinds of beads with appropriate coatings (streptavidin & protein A) and transported to different directions determined by the orientation of the microtubules in a microfluidic device. Bottom-up functionalities, e.g. specificity and throughput of the bio-world were combined with top-down fabrication and handling capabilities of MEMS technology in this research. The resulting hybrid technology, a combination of bottom-up and top-down approaches, has enabled a unique nano-transport system through the assembly of a complex microtubule network. Such a multidirectional, multilayered transport network based on direct manipulation of single microtubules is achieved for the first time. These results show the importance and the potential of hybrid bio/MEMS systems.

本論文は「Nano-scale Transport Driven by Motor Protein along Precisely Assembled Microtubules - A Study for Direct Molecular Handling to Achieve High Sensitivity Detection of Multiple Analytes -(微小管の精密組み立てによる生体分子モータ駆動ナノ輸送―高感度多種分析のための直接分子操作システム―)」と題して英文で書かれており、6章と付録からなる。細胞内での物質輸送を中心的に担うキネシン・微小管系の生体分子モータに着目し、それをマイクロ流体デバイスに組み込むことでナノレベルの輸送システムを構築する方法と、その実証に関して記されている。

第1章は序論であり、本研究の研究背景を述べている。マイクロ流路内の流れによって分子を運ぶ従来の手法では、単一細胞内のタンパク質分析で必要とするような極微量の検体分子の輸送は極めて困難である。この問題に対して、細胞内の物質搬送システムに倣い、生体分子モータによる標的分子の直接搬送を提案し、本論文の目的と研究の意義を提示している。

第2章は、提案したデバイスを作製する際の基礎的な事項に関し、生体分子モータに関する従来の知見と、チップ上での運動観察法を簡単にまとめている。

第3章は、予めキネシンを付加しておいたナノ粒子上に、標的とする分子を選択的に捕獲し、チップ上に固定した微小管の上でそれを望みの方向に搬送するデバイスに関して述べている。個々の作製ステップの検討、ナノ粒子の種類の多様化(シリコン微小構造、プラスチックビーズ、微小油滴、リポソーム等)、微小管をチップ上に固定する際その化学的配向をそろえる手法の検討、などを行った。これらを総合し、上記デバイスの実現と動作に成功した。

第4章では、MEMSピンセットで微小管を個別に扱い、搬送路をガラス基板上に自在に再構成する技術を提案し、その実現方法について述べている。

第5章は、上記の要素技術を総合的に用いて、複数の微小管からなる複雑な搬送経路を造り、キネシン付加ビーズがその上で運動する時の振る舞いについて実験的に明らかにしている。更にこのナノ搬送デバイスの応用を目指して、基礎的な検討を行った結果を述べてある。

第6章は結論であり、本論文で得た成果をまとめ、その意義を論ずるとともに、生体分子モータなどのナノ機能要素をMEMSなどの人工構造中に取り込んだ、ハイブリッドナノシステムに関する研究の進むべき方向を述べている。

以上これを要するに、本論文は、細胞内物質輸送に関わるキネシン・微小管系の生体分子モータに着目し、MEMSピンセットで微小管を個別に扱い、搬送路をガラス基板上に自在に精密組み立てする技術や、キネシンが微小管に沿って運ぶナノ物体の種類や表面修飾を最適化することにより、特定の分子をナノ物体に選択的に付加して搬送する新規な技術を確立して、実際にナノ搬送デバイスの実現と動作確認を行なったもので、電気工学に貢献するところが少なくない。

よって本論文は博士(工学)の学位請求論文として合格と認められる。