Taishi Tonooka

Induced chirality through electromagnetic coupling between chiral molecular layers and plasmonic nanostructures.

We report a new approach for creating chiral plasmonic nanomaterials. A previously unconsidered, far-field mechanism is utilized which enables chirality to be conveyed from a surrounding chiral molecular material to a plasmonic resonance of an achiral metallic nanostructure. Our observations break a currently held preconception that optical properties of plasmonic particles can most effectively be manipulated by molecular materials through near-field effects. We show that far-field electromagnetic coupling between a localized plasmon of a nonchiral nanostructure and a surrounding chiral molecular layer can induce plasmonic chirality much more effectively (by a factor of 10(3)) than previously reported near-field phenomena. We gain insight into the mechanism by comparing our experimental results to a simple electromagnetic model which incorporates a plasmonic object coupled with a chiral molecular medium. Our work offers a new direction for the creation of hybrid molecular plasmonic nanomaterials that display significant chiroptical properties in the visible spectral region.

Lipid bilayers on a picoliter microdroplet array for rapid fluorescence detection of membrane transport.

This paper describes picoliter-sized lipid bilayer chambers and their theoretical model for the rapid detection of membrane transport. To prepare the chambers, semispherical aqueous droplets are patterned on a hydrophilic/hydrophobic substrate and then brought into contact with another aqueous droplet in lipid-dispersed organic solvent, resulting in the formation of the lipid bilayers on the semispherical droplets. The proposed method implements the lipid bilayer chambers with 25-fold higher ratio of lipid membrane area (S) to chamber volume (V) compared to the previous spherical droplet chambers. Using these chambers, we are able to trace the time-course of Ca(2+) influx through α-hemolysin pores by a fluorescent indicator. Moreover, we confirm that the detection time of the substrate transport is inversely proportional to the S/V ratio of the developed chambers, which is consistent with the simulation results based on the developed model. Our chambers and model might be useful for rapid functional analyses of membrane transport phenomena.

pH-Induced Motion Control of Self-Propelled Oil Droplets Using a Hydrolyzable Gemini Cationic Surfactant

Self-propelled motion of micrometer-sized substances has drawn much attention as an autonomous transportation system. One candidate vehicle is a chemically driven micrometer-sized oil droplet. However, to the best of our knowledge, there has been no report of a chemical reaction system controlling the three-dimensional motion of oil droplets underwater. In this study, we developed a molecular system that controlled the self-propelled motion of 4-heptyloxybenzaldehyde oil droplets by using novel gemini cationic surfactants containing carbonate linkages (2G12C). We found that, in emulsions containing sodium hydroxide, the motion time of the self-propelled oil droplets was longer in the presence of 2G12C than in the presence of gemini cationic surfactants without carbonate linkages. Moreover, in 2G12C solution, oil droplets at rest underwent unidirectional, self-propelled motion in a gradient field toward a higher concentration of sodium hydroxide. Even though they stopped within several seconds, they restarted in the same direction. 2G12C was gradually hydrolyzed under basic conditions to produce a pair of the corresponding monomeric surfactants, which exhibit different interfacial properties from 2G12C. The prolonged and restart motion of the oil droplets were explained by the increase in the heterogeneity of the interfacial tension of the oil droplets.

Magnetically responsive microflaps reveal cell membrane boundaries from multiple angles

A microflap system to incline adherent cells in the desired orientation is described. Inclination angles of cell-laden microflaps are precisely controlled by the applied magnetic field, enabling us to observe cell-membrane boundaries from multiple angles. This system is equipped with conventional microscopes, allowing clear focused images of cell-membrane boundaries to be obtained with high magnification.

Rapid and Scalable Preparation of Bacterial Lysates for Cell-Free Gene Expression

Cell-free gene expression systems are emerging as an important platform for a diverse range of synthetic biology and biotechnology applications, including production of robust field-ready biosensors. Here, we combine programmed cellular autolysis with a freeze-thaw or freeze-dry cycle to create a practical, reproducible, and a labor- and cost-effective approach for rapid production of bacterial lysates for cell-free gene expression. Using this method, robust and highly active bacterial cell lysates can be produced without specialized equipment at a wide range of scales, making cell-free gene expression easily and broadly accessible. Moreover, live autolysis strain can be freeze-dried directly and subsequently lysed upon rehydration to produce active lysate. We demonstrate the utility of autolysates for synthetic biology by regulating protein production and degradation, implementing quorum sensing, and showing quantitative protection of linear DNA templates by GamS protein. To allow versatile and sensitive β-galactosidase (LacZ) based readout we produce autolysates with no detectable background LacZ activity and use them to produce sensitive mercury(II) biosensors with LacZ-mediated colorimetric and fluorescent outputs. The autolysis approach can facilitate wider adoption of cell-free technology for cell-free gene expression as well as other synthetic biology and biotechnology applications, such as metabolic engineering, natural product biosynthesis, or proteomics.

Lipid bilayer on a droplet: Formation of lipid bilayers on a droplet array

This paper describes a method for the formation of a lipid bilayer on a droplet-based chamber array. This method enables us to form small chambers sealed with lipid bilayers easily. The lipid membranes are stable and remain for at least 1 day. We successfully observed a transmembrane phenomenon via the lipid membrane on a droplet by fluorescent monitoring. Due to its small volume, subtle changes of molecular amount can be clearly detected. Therefore, we believe that those lipid membrane chambers are useful for a fundamental research of lipid membranes and a high throughput analysis of membrane proteins. By reconstituting membrane proteins (α-hemolysin), we demonstrated that those membranes were lipid bilayers.

Electrical detection of pesticide vapors by biological nanopores with DNA aptamers

This paper describes a vapor detecting system that applies two robust biological elements: A biological nanopore formed in a lipid bilayer and a DNA aptamer. The principle of the sensor is as follows: 1) DNA aptamer selectively captures the target molecules, 2) builds up a molecular complex larger than the nanopore size, and 3) inhibits the ionic current through the nanopore by blocking. We integrated these biological molecules into a previously developed device. A feasibility test was performed using a vapor phase sample, an organophosphorus pesticide, and represented the results demonstrating long-and-deep current blockades with the presence of the omethoate.

Highly packed liposome assemblies toward synthetic tissue

This paper presented an approach for preparation of a highly-packed liposome assembly that implemented lipid bilayer-lipid bilayer contact at the interfaces to mimic a cell-cell connection on living tissues. Cell-sized liposomes were closely packed using our previous technique that allowed monodisperse liposomes arrayed on a substrate. We explored the lipid patterning conditions that would provide a packed structure of the liposomes. By further works, we believe that the assembled structure would be useful for a tissue model.

Rotational chambers on fluidic channels for the repetitive formation of optically observable lipid-bilayer membranes

We develop a device adapted for repetitive formation of horizontal lipid bilayer membranes. This device allows simultaneous optical and electrophysiological measurements of the formed membranes. We integrated a rotational chamber on a fluidic channel separated by a parylene micropore. The rotational motion is designed to form a bilayer repeatedly. This rotational process emulates the conventional painting method, in which a thick lipid-oil layer at a micropore was made thinner by hand work to obtain a bilayer. The bilayer formation with our device was examined optically and electrically. The simultaneous measurement device will be useful for better understanding of bilayer features and membrane protein incorporation.

An inhalation anesthetic device for stereotaxic operation on mouse pups

BACKGROUND Mouse pups are invaluable model animals for understanding the molecular and neural basis underlying behavioral development. Stereotaxic operations with anesthetic control are useful tools in systems neuroscience. However, there are no commercially available anesthetic or stereotaxic devices for mouse pups. Current devices have several problems such as invasive approach for stabilization, poor sanitary control, and less flexibility to combine other surgical apparatuses. NEW METHOD Here, we developed an inhalation anesthetic device equipped with stereotaxic function for mouse pups, by using polydimethylsiloxane (PDMS). PDMS is tolerant to heat and water exposure, and soft enough to cut or make a hole. The anesthetic and the stereotaxic parts were fabricated from the three-dimensional computer-aided design (3D CAD) data obtained from the head of a real mouse pup. RESULTS To confirm its utility, a tracer was injected into the brain. We were able to anesthetize and stabilize pups at once in a non-invasive manner using the PDMS device. The histological staining revealed that tracer injection was successful. Our device was compatible with various types of commercial stereotaxic and anesthetic apparatuses via trimming and tube insertion, respectively. COMPARISON WITH EXISTING METHOD(S) To our knowledge, this is the first report of a device that can stabilize the mouse pups head with the non-invasive manner and functions as an inhalation anesthetic device that can be sterilized. CONCLUSIONS The present fabrication method will provide a handy and functional instrument for stereotaxic operations in animal models at various developmental stages.