Masakuni Sugita

On-chip enucleation of an oocyte by untethered microrobots

We propose a novel on-chip enucleation of an oocyte with zona pellucida by using a combination of untethered microrobots. To achieve enucleation within the closed space of a microfluidic chip, two microrobots, a microknife and a microgripper were integrated into the microfluidic chip. These microrobots were actuated by an external magnetic force produced by permanent magnets placed on the robotic stage. The tip of the microknife was designed by considering the biological geometric feature of an oocyte, i.e. the oocyte has a polar body in maturation stage II. Moreover, the microknife was fabricated by using grayscale lithography, which allows fabrication of three-dimensional microstructures. The microgripper has a gripping function that is independent of the driving mechanism. On-chip enucleation was demonstrated, and the enucleated oocytes are spherical, indicating that the cell membrane of the oocytes remained intact. To confirm successful enucleation using this method, we investigated the viability of oocytes after enucleation. The results show that the production rate, i.e. the ratio between the number of oocytes that reach the blastocyst stage and the number of bovine oocytes after nucleus transfer, is 100%. The technique will contribute to complex cell manipulation such as cell surgery in lab-on-a-chip devices.

Micro-aquatic-farm: On chip stimulation and evaluation system for microorganisms by magnetically driven microtools

In this paper, we propose micro-aquatic-farm, which mimic fluvial environment for an aquatic microorganism. The system provides ideal environment to measure and understand characteristics of the single microorganism. On-chip force sensor is fabricated by assembling layers to neglect the friction issue and it is actuated by permanent magnets, which supply mN order force to stimulate microorganisms. The displacement is magnified by designing beams on the force sensor and the sensor achieved 100 µN resolutions. We succeeded in on-chip stimulation and evaluation of Pleurosira laevis by magnetically driven microtool.

On-chip force sensing by magnetically driven microtool for measurement of stimulant property of P. laevis

In this paper, we newly propose an on-chip force sensing by using a magnetically driven microtool (MMT) equipped with a frame structure for measurement of stimulant property of Pleurosira laevis. The design and fabrication of the force sensing structure with a displacement magnification mechanism based on beam deformation is discussed. Through the basic experiments, the advantages of the proposed layer fabrication technique and the performance of the force sensor are confirmed. The basic characteristics of P. laevis are also confirmed by using the developed MMT.

On-chip manipulation and sensing of microorganisms by magnetically driven microtools with a force sensing structure

In this paper, we introduce an untethered type of magnetically driven microtool (MMT) with a force sensing function to investigate the stimulus characteristics of aquatic microorganisms in a microfluidic chip. The microchip is composed of the proposed MMTs, which have a Si-Ni hybrid structure, a transparent cover made of polydimethylsiloxane (PDMS), a glass substrate, and a microspacer to prevent both friction during the force measurement and damage to the micropattern of the MMT. The fabrication and drive performance of the MMT, which is actuated by permanent magnets from the outside of the microchip, verified in basic experiments. Finally, we succeeded in on-chip manipulation and sensing of microorganisms using this MMT, which we found to be easy to use.

Fabrication of Nanopillar Micropatterns by Hybrid Mask Lithography for Surface-Directed Liquid Flow

This paper presents a novel method for fabricating nanopillar micropatterns for surface-directed liquid flows. It employs hybrid mask lithography, which uses a mask consisting of a combination of a photoresist and nanoparticles in the photolithography process. The nanopillar density is controlled by varying the weight ratio of nanoparticles in the composite mask. Hybrid mask lithography was used to fabricate a surface-directed liquid flow. The effect of the surface-directed liquid flow, which was formed by the air-liquid interface due to nanopillar micropatterns, was evaluated, and the results show that the oscillation of microparticles, when the micro-tool was actuated, was dramatically reduced by using a surface-directed liquid flow. Moreover, the target particle was manipulated individually without non-oscillating ambient particles.

Hybrid mask lithography for fabrication of micro-pattern with nano-pillars

This paper presents the fabrication process of nano-pillar with micro-pattern simultaneously. We proposed the hybrid mask of micro-pattern photo mask and nano-particle mask to obtain the simple fabrication of nano-pillar with micro-pattern. The features of this process are summarized as follows. (1) It is possible to fabricate arbitrary 2D pattern using photolithography. (2) We can get nano-pillar whose size was diffraction-limited by using nano-particle mask. (3) We can control the density of the nano-pillar by changing the weight ratio of nano-particle. (4) We can control the height of the pillar as same the micro-pattern which is the original surface of the substrate.

Free accessible microchannel formed by wide range wettability control using nano-geometric surface

We succeeded in manipulating a microparticle in a free accessible microchannel formed by air-liquid interface by using the on-chip robot. To pattern the stable air-liquid interface in the microfluidic chip, we controlled the wettability of the surface using the nano-geometric surface, which was fabricated by the hybrid mask lithography technique using composite of photoresist and nanoparticle. By adjusting the density of nanopillar coated fluorocarbon or SiO2, the contact angle was controlled from 0 degree to 160 degree. Finally, we demonstrated the on-chip transfer and isolation of the microparticle by the developed free accessible microchannel and the MMT.

Robotic-Investigators for microorganisms in a microfluidic chip

In this paper, we propose 3 degree-of-freedom dual-arm microrobot called Robotic-Investigator (RI) which enables manipulation and force sensing for microorganisms in a microfluidic chip. The untethered RI is actuated by permanent magnets. Therefore, the RI can supply mN order force to stimulate microorganisms with measuring the applied force. To develop the untethered RI with force sensing structure, the layered fabrication technique and on-chip separation mechanism are introduced. Through the basic experiment, we confirmed that the untethered RI with thin structure was developed by proposed fabrication methods without any damage to RI. We also perform the manipulation and force sensing experiment for microorganisms in a microfluidic chip by the developed RI.

3DOF dual-arm microrobots enabling force sensing in a microfluidic chip

In this paper, we propose 3DOF dual-arm microrobots which enabling manipulation and force sensing for a single cell of microorganisms in a microfluidic chip. The untethered microrobots are actuated by permanent magnets which can supply mN order force to stimulate microorganisms and measure the applied force where we want to measure them. The design and fabrication of the untethered microrobots with force sensing structure are discussed. Through the basic experiment, the advantages of the proposed methods and the performance of the 3DOF dual-arm microrobots are confirmed. We also perform the manipulation and force sensing experiment for microorganisms by the developed microrobots.