As a platform for high resolution bioimaging, biochips with a refractive index very close to that of a culture medium (water) are highly demanded. CYTOP (AGC Corporation) is a promising material, since its refractive index (1.34) is almost similar to that of water (1.33). However, high transparency in also the ultraviolet region and high chemical resistance of CYTOP make it difficult to fabricate 3D microstructures for biochips. In this study we present fabrication method of 3D biochips based on CYTOP for cell observation by the molding process using 3D micro/nanostructures fabricated by two-photon polymerization with femtosecond laser.
KEYWORDS: Super resolution, Polymers, Microfluidics, 3D microstructuring, Two photon polymerization, Fabrication, Water, Femtosecond phenomena, Transparency, Refractive index
Development of biochips enabling distortion-free imaging in the microchannel filled with a culture medium (water) is required to investigate behavior of living cells in micro and nano environments. Fluoropolymer CYTOP is a promising material as a platform of biochips for the distortion-free imaging, because the refractive index of CYTOP (1.34) is almost same as 1.33 of water. In this study, we have developed a new 3D fabrication method for CYTOP by two-photon polymerized structures using a femtosecond laser as molds, which will be used for fabrication of micro and nano environment platforms for super-resolution bioimaging.
Laser-induced bubbles can be formed by focusing a conventional nanosecond (ns) laser in a liquid. We recently developed a microfabrication technique (microfabrication using laser-induced bubble (microFLIB)) and applied it to polydimethylsiloxane (PDMS), a thermoset polymer. The technique enabled the rapid fabrication of a high-quality microfluidic channel on a PDMS substrate and selective metallization of the channel via subsequent electroless plating. In addition, we found out that this technique enables true three-dimensional (3D) microfabrication of PDMS so that a hollow microfluidics can be embedded in the polymer substrate. Furthermore, a through hole having high aspect ratio of more than 200 can be fabricated by the single laser scanning. In the experiments, a ns laser beam was focused inside uncured liquid PDMS and was scanned to generate line of laser-induced bubbles. In the microFLIB processing, the shape of the created bubbles was retained in the uncured PDMS for a while; thus, the line of bubbles generated by the laser scanning successfully produced a microfluidic channel on and inside the PDMS substrate after subsequent thermal curing. However, mechanism of this process still remains unclear. Therefore, in this presentation, we will introduce how the microFLIB works in detail and investigate the behaviors of the bubble in the uncured PDMS and explore how the bubble connect to each other during the microFLIB using pump-probe imaging technique.
This conference presentation was prepared for the Laser Applications in Microelectronic and Optoelectronic Manufacturing (LAMOM) XXVIII conference at SPIE LASE, 2023.
Laser-induced bubbles can be formed by focusing a conventional nanosecond (ns) laser in a liquid. We recently developed a microfabrication technique (microfabrication using laser-induced bubble (microFLIB)) and applied it to polydimethylsiloxane (PDMS), a thermoset polymer. The technique enabled the rapid fabrication of a high-quality microfluidic channel on a PDMS substrate and selective metallization of the channel via subsequent electroless plating. In addition, we found out that this technique enables true three-dimensional (3D) microfabrication of PDMS so that a hollow microfluidics can be embedded in the polymer substrate. Furthermore, a through hole having high aspect ratio of more than 200 can be fabricated by the single laser scanning. Therefore, in this presentation, we will introduce how the microFLIB works in detail and demonstrate surface microfabrication of PDMS and 3D microfabrication of hollow microstructures in PDMS. In the experiments, a ns laser beam was focused inside uncured liquid PDMS and was scanned to generate 2D and 3D line of laser-induced bubbles. In the microFLIB processing, the shape of the created bubbles was retained in the uncured PDMS for a while; thus, the line of bubbles generated by the laser scanning successfully produced a microfluidic channel on and inside the PDMS substrate after subsequent thermal curing. The developed microFLIB technique permits the high-speed and high-quality microfabrication of PDMS and can be applied to biochip applications.
Microfluidic chips known as μ-TAS or LoC have become versatile tools in cell research, since functional biochips are able to streamline dynamic observations of various cells. Glass or polymers are generally used as the substrate due to their high transparency, chemical stability and cost-effectiveness. However, these materials are not well suited to the microscopic observation at the fluid boundary due to the refractive index mismatch between the medium and the biochip material. For this reason, we have developed a method of fabricating three-dimensional (3D) microfluidic chips made of a low refractive index fluoric polymer CYTOP. CYTOP has a refractive index of 1.34, a value that is almost equivalent to that of water. This optical property is very important for clear 3D microscopic observations of cell motion near the solid boundary, due to the minimal mismatch between the refractive index values of the medium and the CYTOP substrate. Therefore, CYTOP microfluidics are expected to allow the generation of clear images of unique cell migratory processes near the microfluidic sidewall. Therefore, we established the fabrication procedure involving the use of femtosecond laser direct writing, followed by wet etching and annealing, to create high-quality 3D microfluidics inside a polymer substrate. A microfluidic chip made in this manner enables us to more clearly observe areas near the fluid surface, compared to the observations possible using conventional microfluidic chips.
We present the surface microfabrication of CYTOP, an amorphous UV transparent fluoric polymer, using a conventional pulsed green laser. Fluorine-based polymers have unique characteristics, including high transparency from the deep-UV to the IR wavelengths, chemical stability and low refractive indices, and thus are expected to have a number of industrial applications. However, CYTOP and other fluoric polymers are generally very difficult to microfabricate unless to use high particle beams. Therefore, we have developed a technique for the crack-free surface microfabrication of a CYTOP substrate based on ablation with a pulsed green laser followed by successive wet etching and annealing. In this work, the fundamental surface characteristics of the fabricated area were investigated. After the surface microfabrication of CYTOP, a three dimensional biochip was fabricated using a conventional PDMS bonding technique to demonstrate that a CYTOP biochip is superior to conventional transparent biochips with regard to the microscopic observation of cell motion.
We demonstrate fabrication of microfluidic chips integrated with optical waveguide embedded in a photostructurable
glass for high-sensitive biochemical liquid analysis using a femtosecond laser. Femtosecond laser direct writing followed
by annealing and successive wet etching in hydrofluoric acid solution resulted in the rapid manufacturing of microfluidic
chips for the biochemical liquid concentration assay. By covering the internal wall of the microfluidic channel in the
glass with low refractive index polymer, interaction between the liquid and incident analyzing light is enhanced.
Therefore, the microfluidic chip enables us to analyze the low concentrations down to 7.5 mM of protein in bovine
serum albumin. Such microfluidic chip realizes the efficient and high-sensitive concentration analysis of biochemical
liquids at early stages of biochemical reactions.
We demonstrate three-dimensional (3D) microstructuring inside glass by ultrafast laser to fabricate microfluidic chips
integrated with some functional microcomponents such as optical attenuators and optical waveguides. The fabricated
microchips are applied to understand phenomena and functions of microorganisms and cyanobacteria. Ultrafast laser
irradiation followed by thermal treatment and wet etching in dilute hydrofluoric acid solution resulted in fabrication of
3D microfludic structures embedded in a photosensitive glass. The embedded microfludic structures enabled us to easily
and efficiently observe Phormidium gliding to the seedling root, which accelerates growth of the vegetable. In addition,
integration of optical attenuators and optical waveguides into the microfluidic structures clarified the mechanism of the
gliding movement of Phormidium. We termed such integrated microchips nanoaquariums, realizing the highly efficient
and functional observation and analysis of various microorganisms.
We demonstrate to fabricate microfluidic chips integrated with some functional microcomponents such as optical
attenuators and optical waveguides by femtosecond laser direct writing for understanding phenomena and functions of
microorganisms. Femtosecond laser irradiation followed by annealing and wet etching in dilute hydrofluoric acid
solution resulted in fabrication of three-dimensional microfludic structures embedded in a photosensitive glass. The
embedded microfludic structures enabled us to easily and efficiently observe Phormidium gliding to the seedling root,
which accelerates growth of the vegetable. In addition, integration of optical attenuators and optical waveguides into the
microfluidic structures clarified the mechanism of the gliding movement of Phormidium. We termed such integrated
microchips nanoaquariums, realizing the highly efficient and functional observation and analysis of various
microorganisms.
We demonstrate fabrication of microfluidic chips integrated with different functional elements such as optical filters and
optical waveguide for mechanism study of gliding movement of Phormidium to a seedling root using a femtosecond (fs)
laser. Fs laser direct writing followed by annealing and successive wet etching in dilute hydrofluoric (HF) acid solution
resulted in formation of three dimensional (3D) hollow microstructures embedded in a photostructurable glass. The
embedded microfludic structures enabled us to easily and efficiently observe Phormidium gliding to the seedling root,
which accelerates growth of the seedling. In addition, fabrication of optical filter and optical waveguide integrated with
the microfluidic structures in the microchip clarified the mechanism of the gliding movement. Such microchips, referred
to as a nano-aquarium, realize the efficient and highly functional observation and analysis of the gliding movement of
Phormidium.
A microchip made of UV transparent polymer (CYTOP) that can perform selective cell culture has been fabricated by F2
laser surface modification. The refractive index of CYTOP is almost the same as that of culture medium, which is
essential for three dimensional (3D) observation of cells. The F2 laser modification of CYTOP achieves hydrophilicity
only on the laser irradiated area with little deterioration of the optical properties and surface smoothness. After the laser
modification, HeLa cells were successfully cultured and strongly adhered only on the modified area of CYTOP. The
cells patterned on CYTOP were applied for clear 3D observation using an optical microscope in phase contrast mode.
We demonstrate the fabrication of three-dimensional (3D) hollow microstructures embedded in photostructurable glass by a nonlinear multiphoton absorption process using a femtosecond (fs) laser. Fs laser direct writing followed by annealing and successive wet etching in dilute hydrofluoric (HF) acid solution resulted in the rapid manufacturing of microchips with 3-D hollow microstructures for the dynamic observation of living microorganisms in fresh water. The embedded microchannel structure enables us to analyze the continuous motion of Euglena gracilis and Cryptomonas. Such microchips, referred to as nano-aquariums realize the efficient and highly functional observation of microorganisms.
Three-dimensional (3D) microfabrication of photosensitive glass by femtosecond (fs) laser direct writing is demonstrated
for manufacture of microchips for biomedical applications. The fs laser direct writing followed by annealing and
successive wet etching can fabricate the hollow microstructures, achieving a vareiety of microfluidic components and
microoptical components in a glass chip. One of the interesting and important applications of the 3D microfluidic
structures fabricated by the present technique is inspection of living microorganisms. The microchips used for this
application are referred to as nanoaquarium. Furthermore, the optical waveguide is written inside the glass by the fs
laser direct writing without the annealing and the successive etching. It is revealed that integration of the microfluidic
and microoptical components with the optical waveguides in a single glass chip is of great use for biochemical analysis
and medical inspection based on optical sensing.
We demonstrate the fabrication of three-dimensional (3-D) hollow microstructures embedded in photostructurable glass
by a femtosecond (fs) laser direct writing. Fs laser direct writing followed by annealing and successive wet etching in
dilute hydrofluoric (HF) acid solution resulted in the rapid manufacturing of microchips with 3-D hollow microstructures
for the dynamic observation of living microorganisms in fresh water. The embedded microchannel structure enables us
to analyze the continuous motion of Euglena gracilis. A microchamber with a movable microneedle demonstrates its
ability for the elucidation of the information transmission process in Pleurosira laevis. Such microchips, referred to as
nano-aquariums realize the efficient and highly functional observation of microorganisms.
We demonstrate the fabrication of three-dimensional (3D) hollow microstructures embedded in photostructurable glass
by a nonlinear multiphoton absorption process using a femtosecond (fs) laser. Fs laser direct writing followed by
annealing and successive wet etching in dilute hydrofluoric (HF) acid solution resulted in the rapid manufacturing of
microchips with 3-D hollow microstructures for the dynamic observation of living microorganisms in fresh water. The
embedded microchannel structure enables us to analyze the continuous motion of Euglena gracilis and Dinoflagellate.
Such microchips, referred to as nano-aquariums realize the efficient and highly functional observation of
microorganisms.
Three-dimensional microfabrication of photostructurable glass by femtosecond (fs) laser direct writing is demonstrated
for manufacture of photonic biochips. The fs laser direct writing followed by annealing and successive wet etching can
fabricate the hollow microstructures, achieving a vareiety of microfluidic components and microoptical components in a
glass chip. The optical waveguide written by the fs laser direct writing without the annealing and the successive etching
can be integrated into these microfluidic and microoptical circuits. Such an integrated microchip is of great use for
biochemical analysis and medical inspection based on photonic sensing. Furthermore, as one of the interesting and
important applications, the microchips fabricated by the present technique are applied for inspection of living
microorganisms.
The laser-induced plasma-assisted ablation (LIPAA) process developed by our group, in which a single conventional pulsed laser is only used, makes it possible to perform high-quality and high-speed glass microfabrication. Up to the present, this process has been widely applied for micromachining of various transparent hard and soft materials. In this process, the laser beam first passes through the glass substrate since the laser beam has no absorption by the substrate. Then, the transmitted beam is absorbed by a solid target (typically a metal), located behind the substrate so that the target is ablated, resulting in plasma generation. Due to the interaction of the laser beam and the laser-induced plasma, significant ablation takes place at the rear surface of the substrate. Recently, we have developed the proto-type LIPAA system using a second harmonic of diode pumped Q-switched Nd:YAG laser for the practical use. In this paper, we demonstrate micromachining, crack-free marking and color marking of glass materials. Additionally, selective metallization of glass and polyimide by the LIPAA process followed by metal chemical-plating is investigated. A possible mechanism of LIPAA is also discussed based on the results from double pulse irradiation using near-IR fs laser, transient absorption measurement and plasma-conductivity measurement.
The laser-induced plasma-assisted ablation (LIPAA) process developed by our group, in which a single conventional pulsed laser is only used, makes it possible to perform high-quality and high-speed glass microfabrication. Up to the present, this process has been widely applied for micromachining of various transparent hard and soft materials. In this process, a laser beam is first directed to a glass substrate placed in vacuum or air. The laser beam passes through the substrate since the wavelength of the laser beam must have no absorption by the substrate for the LIPAA process. The transmitted laser beam is absorbed by a solid target (typically a metal), located behind the substrate. The target is then ablated, resulting in plasma generation. Due to the interaction of the laser beam and the laser-induced plasma, significant ablation takes place at the rear surface of the substrate. Recently, we have developed the proto-type LIPAA system using a second harmonic of diode pumped Q-switched Nd:YAG laser for the practical use. In this paper, we will demonstrate micromachining, crack-free marking, color marking and dicing of glass materials. Additionally, selective metallization of glass and polyimide by the LIPAA process followed by metal chemical-plating is investigated. The discussion includes mechanism and practical applications in micro-electronics industry of the LIPAA process.
Laser-induced plasma-assisted ablation (LIPAA) process developed for glass materials has been applied for micromachining of a variety of transparent hard and soft materials. We have developed the proto-type LIPAA system using a second harmonic of diode pumped Q-switched Nd:YAG laser for the practical use. In this paper, micromachining and scribing of glass and sapphire is demonstrated using the developed system. Additionally, another application such as selective metallization of glass and polyimide with successive metal plating process is investigated. However, mechanism of this process is complex and still remains unknown. To have a better understanding of this process, double-pulse irradiation of a near-IR femtosecond (fs) laser with a delay time is also investigated. A possible mechanism is discussed based on the obtained results.
The specific deposition and atomic redistribution with use of the phase-shifted laser were examined. Phase-shifting is a unique technique to realize the abrupt intensity change in a very narrow region of focused laser beam. This technique is applied to an ultra-short laser pulse to study the interaction between particles removed from very closely separated ablation spots in focal area. Using the phase- shifted focused laser beam, atomically nonuniform deposition was observed in a femtosecond laser ablation of Cu:Zn alloy target. Atomic concentration ratio of Cu and Zn was obtained by the measurement of energy dispersive X-ray analyzer.
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