Keisuke Takada

A new biosensing by Dielectric Dispersion Analysis of interaction between lipid membrane of liposome and target biomolecules up to 20 GHz range

This paper reports, firstly, that it was newly found by our specific Dielectric Dispersion Analysis (DDA) that plural type of bound water with different relaxation frequencies exist in a suspension of liposome of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). Secondly, we observe that the interaction between the liposome and a specific target biomolecule depends on the type of target molecules (we used a protein of carbonic anhydrase from bovine (CAB) and/or cholesterol, respectively), leading to the change in dielectric relaxation at different relaxation frequency of bound water. It is finally concluded that the developed DDA method allows us to acquire the quantitative information with respect to bound water to liposomes, which gives novel physical insight on interaction between liposomes and target biomolecules (especially, protein).

Detection of Amyloid Beta Fibril Growth by Liposome-Immobilized Micro-Cantilever With NiCr Thin-Film Strain Gauge

A micro-cantilever with thermally stable strain gauge of a NiCr thin film and a self-assembled monolayer was fabricated by surface micromachining. Liposomes, as biosensing molecules, were immobilized on its surface. To prevent evaporation of the water solvent, a reservoir made of polydimethylsiloxane around the micro-cantilever was fabricated and covered with a glass plate. The resistance of the strain gauge increases with time in the amyloid beta (Aβ) solution, because the cantilever is bent downward caused by interaction between Aβ and liposomes immobilized on the micro-cantilever surface. Moreover, resistance change depends on the interaction ability of Aβ proteins with liposomes. It is therefore indicated that the fabricated static mode micro-cantilever sensor with immobilized liposomes can be used to detect Aβ proteins.

Detection of interaction between biological proteins and immobilized liposomes by a micro-cantilever with NiCr thin film strain gauge

A micro-cantilever with thermally stable strain gauge of a NiCr thin film and a self-assembled monolayer (SAM) was fabricated by surface micromachining and liposomes, as biosensing molecules, were immobilized on its surface. The resistance of the strain gauge increases with time in a biological protein (carbonic anhydrase from bovine: CAB) solution because the cantilever is bent downward by surface adsorption of CAB molecules. On the other hand, the resistance changes differently with the growth of another type protein, amyloid beta (Aβ) fibrils. It is therefore demonstrated that the fabricated static mode micro-cantilever sensor can detect the interaction between the liposomes and the biological proteins.

Bioassay of proteins in stable solution state using a novel cantilever-based liposome biosensor

We have developed a micro-cantilever biosensor with embedded NiCr thin film strain gauge. Meanwhile, DPPC liposomes have been selected as sensing biomolecules to be immobilized on the surface of the micro-cantilever. The liposome-protein interaction was detected by measuring the resistance change rate of the strain gauge. In this work, a PDMS-based sealed reservoir structure was newly fabricated and added on the sensor to keep the target solution stable, where a long-time stable detection was achieved. The resistance of the cantilever-based biosensor increased with time in lysozyme or carbonic anhydrase from bovine (CAB) aqueous solution, and the characteristic of chronological resistance change varied with the concentration and kind of the proteins. It is expected that the micro-cantilever sensor with droplet-sealing structure can be used for bioassay of various proteins in future.

Sensing of biomolecular motion of liposome and target protein, and their interaction by dielectric dispersion analysis for 100–1000 MHz range

We have newly examined biochemical interaction between liposome and target proteins by Dielectric Dispersion Analysis (DDA), especially for frequency range of 100-1000 MHz to consider molecular dynamics of the coexistence of the liposome membrane and target protein. Finally, we observed several new dielectric relaxations by the interaction in the low frequency range, thereafter we newly found and considered that some interaction manners between the each protein against liposome would exist and be different at least qualitatively for the two proteins because the manner of change in relaxation was different between each other. On the other hand, we have detected successfully much smaller dielectric relaxation even in narrower frequency range, thus also able to detect the change with much smaller concentration of protein than the previous works.

Bioassay of Target Proteins Using A NiCr Strain Gauge-Cantilever Liposome Biosensor with Droplet-Sealed Structure

We have developed Si micro-cantilever biosensor with embedded NiCr thin film strain gauge and DPPC liposome immobilized on its surface. This time, a PDMS-based sealed reservoir structure was newly fabricated and added on the sensor to keep the protein solution stable, where a long-time stable detection was achieved. The resistance of the cantilever sensor increased with time in carbonic anhydrase from bovine (CAB) or lysozyme aqueous solution, and the characteristic of chronological resistance change varied with the kind and concentration of proteins. We expect that this micro-cantilever sensor with droplet sealed structure becomes an effective device for bioassay of proteins.

A Dielectric Dispersion Analysis using microwave bio-microsensor for droplet of liposome suspension with target biomolecules

This paper reports, firstly, that it was newly found by our specific Dielectric Dispersion Analysis (DDA) that plural type of bound water with different relaxation frequencies exist in a suspension of liposome of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). Secondly, we observe that the interaction between the liposome and a specific target biomolecule depends on the type of target molecules (protein of carbonic anhydrase from bovine (CAB) and/or cholesterol, respectively), leading to the change in dielectric relaxation at different relaxation frequency of bound water.

A new dielectric dispersion analysis using microwave bio-microsensor for minute droplet of liposome suspension with target biomolecules by S-parameter method

In this work, a microwave bio-microsensor of coplanar waveguide (CPW) interdigital capacitor (IDC) with droplet of about 0.3 μL of liposome suspension with target biomolecules is newly considered and applied for realizing the evaluation of the permittivity of small target droplet by S-parameter method, similar to Dielectric Dispersion Analysis (DDA) by an open-ended probe method. Finally, we have successfully detected by the IDC microsensor the interaction between the liposome and target protein of carbonic anhydrase from bovine (CAB), which is intrinsically the same as that observed by the conventional open-ended probe method. It is significantly important that the IDC method improves the sensitivity, defined as (Δεr/εr)/(solution volume), by about 300 times from the conventional open-ended probe method.

A new intact immobilization of liposome as sensing bio nano-particle on oxidized metal electrode surface

We newly report that the intactness of liposome on the electrode surfaces of Pt and Au metals could be significantly improved on thermally oxidized metal surfaces, compared to those with non-oxidization or even with a SAM treatment. It is especially noted that the sub-gel phase of lipid membrane of the liposome on the oxidized Pt surface is much more suppressed than the case of non-oxidized, indicating that the fluidity of intact liposome greatly improves, as the sub-gel phase then crystalline phase will increase when the intact liposome loses its closed spherical shape. Finally, the retention time of the intact liposome on the oxidized surface becomes improved to about one day. These results indicate that the oxidation treatment made the metal surface hydrophilic enough to keep the stable and intact liposome. We think that the retention time up to one day is good enough for practical biosensor applications using intact liposome to improve device performance and its reliabilities.