Masashi Ikeuchi

Development of pressure-driven micro active catheter using membrane micro emboss following excimer laser ablation (MeME-X) process

The worlds smallest micro active catheter of φ∼300µm was successfully fabricated using membrane micro emboss following excimer laser ablation (MeME-X) process. The catheter has a one-sided hollow bellows at the tip made of thin polymer membrane. The bellows is composed of a series of folded micro-chambers and microchannels connecting the micro-chambers. The folded-chambers expand on one side by increasing inner water pressure using a syringe, thus the whole bellows bends toward one direction within 0 to 180 degree. This micro active catheter should be useful for safe intravascular surgery in narrow and complicated blood vessels. Moreover, the nonelectrical actuation mechanism of this catheter can be widely applied to micro-actuators and micro-robots which need high safety.

Electrospray deposition and direct patterning of polylactic acid nanofibrous microcapsules for tissue engineering.

Electrospun nanofibers composed of biodegradable polymers are attractive candidates for cell culture scaffolds in tissue engineering. Their fine-meshed structures, resembling natural extracellular matrices, effectively interact with cell surfaces and promote cell proliferation. The application of electrospinning, however, is limited to two-dimensional (2D) or single tube-like scaffolds, and the fabrication of arbitrary three-dimensional (3D) scaffolds from electrospun nanofibers is still very difficult due to the fibers’ continuous and entangled form. To address this issue, in this paper, we describe the use of phase-separation-assisted electrospray and electrostatic focusing to perform continuous direct 3D patterning of nanofibrous microcapsules of biodegradable polylactic acid (PLA). These microcapsules exhibit fiber-particle duality because they are composed of nanofibers suitable for cell attachment while also being easy to handle as particles for direct 3D patterning. By varying the flow rate of the polymer solution and the humidity of the electrospray atmosphere during electrospraying, the diameter of the microcapsule and its surface porosity can be controlled. The utility of the direct-patterning process is demonstrated by fabricating high-aspect-ratio microscaffolds and subsequent cell cultures. The nanofibrous and hollow structure of the microcapsules combined with the direct 3D patterning process offers a new approach for fabricating tailor-made scaffolds for regenerative medicine.

“Membrane micro emboss following excimer laser ablation (MeME-X) process” for pressure-driven micro active catheter

We have developed MeME-X process to break-through the conventional miniaturization limit of the pressure-driven micro active catheter by two orders of magnitude. The worlds smallest micro active catheter of r~200mum was successfully fabricated using MeME-X process. The catheter was made of thin biocompatible polymer membrane and actuated just by water pressure for safe intravascular surgery.

Spheroid Formation and Evaluation of Hepatic Cells in a Three-Dimensional Culture Device

In drug discovery, it is very important to evaluate liver cells within an organism. Compared to 2D culture methods, the development of 3D culture techniques for liver cells has been successful in maintaining long-term liver functionality with the formation of a hepatic-specific structure. The key to performing drug testing is the establishment of a stable in vitro evaluation system. In this article, we report a Tapered Stencil for Cluster Culture (TASCL) device developed to create liver spheroids in vitro. The TASCL device will be applied as a toxicity evaluation system for drug discovery. The TASCL device was created with an overall size of 10 mm × 10 mm, containing 400 microwells with a top aperture (500 µm × 500 µm) and a bottom aperture (300 µm diameter circular) per microwell. We evaluated the formation, recovery, and size of HepG2 spheroids in the TASCL device. The formation and recovery were both nearly 100%, and the size of the HepG2 spheroids increased with an increase in the initial cell seeding density. There were no significant differences in the sizes of the spheroids among the microwells. In addition, the HepG2 spheroids obtained using the TASCL device were alive and produced albumin. The morphology of the HepG2 spheroids was investigated using FE-SEM. The spheroids in the microwells exhibited perfectly spherical aggregation. In this report, by adjusting the size of the microwells of the TASCL device, uniform HepG2 spheroids were created, and the device facilitated more precise measurements of the liver function per HepG2 spheroid. Our TASCL device will be useful for application as a toxicity evaluation system for drug testing.

Multifunctional optically driven microrobot for realtime 3D bio-manipulation and imaging

We have been working on optically driven microrobots capable of single cell manipulation and real-time reaction force sensing in liquid. In this report, we have developed three key technologies to realize 3D manipulation, imaging and sensing of a single living cell. (1) Fluorescent microrobot for long time imaging. (2) Surface functionalization of the robot for effective capture of cells. (3) 3D manipulation and imaging system for realtime observation. These results should lead to integrated micro robotic system that can remotely manipulate living cells in liquid to study cell-cell interactions, cellular response to mechanical stimulation, and to assemble cells into 3D structures for tissue engineering.

Magnetic micro actuator with neutral buoyancy and 3D fabrication of cell size magnetized structure

We have developed two technologies for 3D magnetic microstructures, with a wide size range between 5μm to 2mm. The first technology enables us to obtain density controlled 3D magnetic microstructures. The size of this structure is approximately 500μm. In this scale, controlling density is vital for magnetic micro actuators, because the effect of gravity is strong. To adjust density, we developed the worlds first “density controllable magnetically photocurable (DMPC) polymer.” The DMPC polymer is a mixture of hollow microcapsules (density, 0.03 g/cm3), magnetic particles, and photocurable polymer. We can obtain desired relative density between 0.5 to 1.7 by adjusting the concentration of microcapsules. In addition, we succeeded in 3D velocity control of a screw-type magnetic micro actuator with neutral buoyancy in water. The delay time was 32msec, which is smaller than video rate. In addition, the actuator possessed 6 degrees of freedom. The second technology realized a 5μm magnetic micro actuator, which is a combination of a 3D transparent structure and 2D magnetic structure. Various photocurable polymers can be applied as the 2D structure in this process, although we used magnetically photocurable polymer in this report. Furthermore, we have succeeded in driving a ferromagnetic micro actuator, whose diameter is as small as 1μm. These two fabrication processes will become key technologies in both the medical field and the field of life sciences, because they can supply a wide variety of 3D micro structures with small effort.

Nanofibrous Surface Patterning using Nano-Meshed Microcapsules Induced by Phase Seperation Assisted Electrospray

This is the first report on a nanofibrous surface patterning process using nano-meshed polymer microcapsules for biological applications. The nano-meshed microcapsules were formed using a method named phase separation assisted electrospray. Features of the nano-meshed microcapsules were found to be tunable by adjusting process conditions. Contrary to non-woven mats formed of continuous nanofibers via conventional electrospinning, nano-meshed microcapsule reported herein enabled a single step nanofibrous surface patterning by electrostatic focusing following the microcapsule formation. The biodegradable polylactic acid made microcapsules were patterned onto a glass substrate with up to 500¿m resolution and human hepatocyte cells were cultured on the patterned areas to confirm biocompatibility of the microcapsule.

Combinatorial differentiation induction of embrionic bodies in "PASCL (Pneumatically Actuated Spheroids Culture Lab-on-chip)"

This paper reports the fabrication, combinatorial differentiation and analysis of embryonic bodies (EBs) in one device called “PASCL (Pneumatically Actuated Spheroids Culture Lab-on-chip)”. We demonstrated EBs formation from human adipose derived stem cells (ADSC), differentiation into adipocytes and osteoblasts and staining for differentiation monitoring all in one PASCL.

Density controllable photocurable polymer for three-dimensional magnetic microstructures with neutral buoyancy

We have developed a density controllable magnetically modified photocurable polymer. Using this developed polymer and microstereolithography, we fabricated three-dimensional magnetic microstructures with neutral buoyancy. This polymer is composed of photocurable polymer, magnetic particles, a viscosity increasing agency, and hollow microcapsules. Because the density of the hollow microcapsules is low, we can adjust the density of the polymer by changing the concentration of the microcapsules. Furthermore, we constructed a mathematical model for the density and magnetic properties of the density controllable magnetically modified photocurable polymer. We expect our developed polymer will become a key material for achieving three-dimensional controllable magnetic micromachines.

Development of New Biochemical IC Chip-Set for Real-Time PCR

We have developed new biochemical IC chip-set which realizes real-time PCR in the finger top size. Unlike conventional ¿-TAS approach, our chip-set has great versatility and portability since all chips are designed in standardized micro-architecture and the fabrication process can be combined with conventional MEMS processes. Combining this chip-set with existing Biochemical IC chip-set family, researchers can easily develop order-made palm-top system for proteomics research, on-site diagnosis, and tailor-made medicine.