High Throughput,Polymeric Aqueous Two‐Phase Printing of Tumor Spheroids

This paper presents a new 3D culture microtechnology for high throughput production of tumor spheroids and validates its utility for screening anti‐cancer drugs. Two immiscible polymeric aqueous solutions are used and a submicroliter drop of the “patterning” phase containing cells is microprinted into a bath of the “immersion” phase. Selecting proper formulations of biphasic systems using a panel of biocompatible polymers results in the formation of a round drop that confines cells to facilitate spontaneous formation of a spheroid without any external stimuli. Adapting this approach to robotic tools enables straightforward generation and maintenance of spheroids of well‐defined size in standard microwell plates and biochemical analysis of spheroids in situ, which is not possible with existing techniques for spheroid culture. To enable high throughput screening, a phase diagram is established to identify minimum cell densities within specific volumes of the patterning drop to result in a single spheroid. Spheroids show normal growth over long‐term incubation and dose‐dependent decrease in cellular viability when treated with drug compounds, but present significant resistance compared to monolayer cultures. The unprecedented ease of implementing this microtechnology and its robust performance will benefit high throughput studies of drug screening against cancer cells with physiologically relevant 3D tumor models.     

Rehydration of Polymeric,Aqueous, Biphasic System Facilitates High Throughput Cell Exclusion Patterning for Cell Migration Studies

This paper describes a cell‐exclusion patterning method facilitated by a polymeric aqueous two‐phase system. The immersion aqueous phase (polyethylene glycol) containing cells rehydrates a dried disk of the denser phase (dextran) on the substrate to form a dextran droplet. With the right properties of the phase‐forming polymers, the rehydrating droplet remains immiscible with the immersion phase. Proper formulation of the two‐phase system ensures that the interfacial tension between the rehydrating droplet and the surrounding aqueous phase prevents cells from crossing the interface so that cells only adhere to the regions of the substrate around the dextran phase droplet. Washing out the patterning two‐phase reagents reveals a cell monolayer containing a well‐defined circular gap that serves as the migration niche for cells of the monolayer. Migration of cells into the cell‐excluded area is readily visualized and quantified over time. A 96‐well plate format of this “gap healing” migration assay demonstrates the ability to detect inhibition of cell migration by known cytoskeleton targeting agents. This straightforward method, which only requires a conventional liquid handler and readily prepared polymer solutions, opens new opportunities for high throughput cell migration assays.     

Rapid Self‐Assembly of Macroscale Tissue Constructs at Biphasic Aqueous Interfaces

An entirely new approach to tissue engineering is presented that uses the interfacial forces between aqueous solutions of phase‐separating polymers to confine cells and promote their assembly into interconnected, macroscopic tissue constructs. This simple and inexpensive general procedure creates free‐standing, centimeter‐scale constructs from cell suspensions at the interface between poly(ethylene glycol) and dextran aqueous two‐phase systems in as little as 2 h. Using this method, skin constructs are produced that integrate with decellularized dermal matrices, on which they differentiate and stratify into skin equivalents. It is demonstrated that the constructs produced by this method have appropriate integrity and mechanical properties for use as in vitro tissue models.     

Two‐Phase Emulgels for Direct Ink Writing of Skin‐Bearing Architectures

Direct ink writing (DIW) provides programmable and customizable platforms to engineer hierarchically organized constructs. However, one‐step, facile synthesis of such architectures via DIW has been challenging. This study introduces inks based on two‐phase emulgels for direct printing and in situ formation of protecting layers enveloping multicomponent cores, mimicking skin‐bearing biological systems. The emulgel consists of a Pickering emulsion with an organic, internal phase containing poly(lactic acid) stabilized by chitin/cellulose nanofibers and a continuous, cross‐linkable hydrogel containing cellulose nanofibers and any of the given solid particles. The shear during ink extrusion through nozzles of low surface energy facilitates the generation of the enveloped structures via fast and spontaneous phase separation of the emulgel. The skin‐bearing architectures enable control of mass transport as a novel configuration for cargo release. As a demonstration, a hydrophilic molecule is loaded in the hydrogel, which is released through the core and skin, enabling regulation of diffusion and permeation phenomena. This 3D‐printed functional material allows independent control of strength owing to the hierarchical construction. The new method of fabrication is proposed as a simple way to achieve protection, regulation, and sensation, taking the example of the functions of skins and cuticles, which are ubiquitous in nature.     

Dual Phase‐Controlled Synthesis of Uniform Lanthanide‐Doped NaGdF4 Upconversion Nanocrystals Via an OA/Ionic Liquid Two‐Phase System for In Vivo Dual‐Modality Imaging

A novel OA/ionic liquid two‐phase system combining the merits of thermal decomposition method, the IL‐based strategy, and the two‐phase approach is introduced to synthesize high‐quality lanthanide‐doped NaGdF₄ upconversion nanocrystals with different crystal‐phases in OA‐phase and IL‐phase through a one‐step controllable reaction. Oil‐dispersible cubic‐phase NaGdF₄:Yb, Er (Ho, Tm) nanocrystals with ultra‐small size (～5 nm) and monodispersity are obtained in the OA phase of the two‐phase system via an IL‐based reaction. More importantly, water‐soluble hexagonal‐phase NaGdF₄:Yb, Er nanocrystals are obtained in the same system simply by adopting an extremely facile method to complete the dual phase‐transition (crystal‐phase transition and OA‐phase to IL‐phase transition) simultaneously. The synthesized lanthanide‐doped NaGdF₄ upconversion nanocrystals are effective for dual‐mode UCL imaging and CT imaging in vivo.     

A Microfluidic Tool for Fine‐Tuning Motion of Soft Micromotors

Microfluidics is an ideal tool for the design of self‐assembled micromotors. It allows for easy change of solutions, catalysts, and flow rates, which affect shape, structure, and motion of the resulting micromotors. A microfluidic tool generating aqueous‐two‐phase‐separating droplets of UV‐polymerizable poly(ethylene glycol)diacrylate (PEGDA) and an inert phase, salt, or polysaccharide, is utilized to fabricate asymmetric microbeads. Different molecular weights and branching of polysaccharides are used to study the effect on shape, surface roughness, and motion of the particles. The molecular weight of the polysaccharide determines the roughness of the motors inner surface. Smooth openings are obtained by low molecular weight dextran, while high surface roughness is obtained with a high molecular weight branched polysaccharide. Since roughness plays an important role in bubble pinning, it influences both speed and trajectory. Increasing speeds are obtained with increasing roughness and trajectories ranging from linear, circular to tumble‐and‐run depending on the nature of bubble pinning. This microfluidic tool allows for fine‐tuning shape, structure, and motion by easy change of solutions, catalysts, and flow rates.     

Study on Two‐Coil and Four‐Coil Wireless Power Transfer Systems Using Z‐Parameter Approach

A wireless power transfer (WPT) system is usually classified as being of either a two‐coil or four‐coil type. It is known that two‐coil WPT systems are suitable for short‐range transmissions, whereas four‐coil WPT systems are suitable for mid‐range transmissions. However, this paper reveals that the two aforementioned types of WPT system are alike in terms of their performance and characteristics, differing only when it comes to their matching‐network configurations. In this paper, we first find the optimum load and source conditions using Z‐parameters. Then, we estimate the maximum power transfer efficiency under the optimum load and source conditions, and we describe how to configure the matching networks pertaining to both types of WPT system for the given optimum load and source conditions. The two types of WPT system show the same performance with respect to the coupling coefficient and load impedance. Further, they also demonstrate an identical performance in the two cases considered in this paper, that is, a strong‐coupled case and a weak‐coupled case.     

Synthesis of Two‐Dimensional Perovskite by Inverse Temperature Crystallization and Studies of Exciton States by Two‐Photon Excitation Spectroscopy

Two‐dimensional (2D) organic–inorganic hybrid perovskites (OIHPs), a natural multiple‐quantum‐well structure with quasi‐2D electronic properties, have recently emerged as a promising class of semiconducting materials for photovoltaic and optoelectronic applications. However, facile synthesis of high‐quality 2D OIHPs single crystals is still lacking. The layer dependence of the exciton binding energy of (C₄H₉NH₃)₂PbI₄ (C4PI), a widely studied 2D OIHP, is still debated. Herein, a novel synthesis technique based on inverse temperature crystallization in a binary‐solvent system is used to prepare 2D OIHPs and a systematic study of excitonic states of the synthesized 2D OIHPs by two‐photon excitation (TPE) spectroscopy is conducted. The obtained TPE spectra indicate that the exciton binding energies are similar for C4PI nanosheets and bulk crystals with different number of layers, most likely due to the intrinsically weak interlayer coupling. Further, the dark excitonic 2p states of (C₆H₅(CH₂)₂NH₃)₂PbI₄ (PEPI) and C4PI are also observed by TPE spectroscopy. The results provide a novel synthesis protocol and insight into exciton properties of 2D OIHPs.     

A New Series of Quadrupolar Type Two‐Photon Absorption Chromophores Bearing 11, 12‐Dibutoxydibenzo[a,c]‐phenazine Bridged Amines; Their Applications in Two‐Photon Fluorescence Imaging and Two‐Photon Photodynamic Therapy

A new series of quadrupolar type two‐photon absorption (2PA) chromophores 3 – 9 bearing a core arylamine‐[a,c]phenazine‐arylamine motif are synthesized in high yields. Palladium‐catalyzed Stille coupling and C? N coupling reactions are utilized to prepare target chromophores. Detailed characterization and systematic studies of these molecules, including absorption and fluorescence emission, are conducted. These compounds are found to exhibit very large 2PA cross section values, for example, ～7000 GM at 800 nm for 8 in toluene. Two‐photon‐induced fluorescence imaging is successfully demonstrated in vitro using compound‐ 8 ‐encapsulated silica nanoparticles with excellent bio‐compatibility. In combination with the capability of both one‐ and two‐photon singlet‐oxygen sensitizations, this nanocomposite demonstrates its promising potential in dual functionality toward two‐photon fluorescence imaging and two‐photon photodynamic therapy.     

Near‐Field Lithography by Two‐Photon Induced Photocleavage of Organic Monolayers

We prove that the enhanced electromagnetic near‐field around metallic nanostructures can be used for localized two‐photon induced activation of surfaces, obtaining a defined chemical pattern with nanometric resolution. Gold nanoparticles (Au‐NP) are deposited on glass slides that were modified with a polysiloxane layer containing a nitroveratrylcarbonyl (NVoc) photoremovable group. Upon illumination with a femtosecond laser, the NVoc entity is removed. Due to the electromagnetic field enhancement of the nanoparticles, the threshold of this process is lowered in the nm‐scale vicinity of the metal structures. Upon cleavage, an amine functional group is released, which can be used to site‐selectively bind species with complementary chemical functionality on the surface. This method can be utilized for sub‐wavelength chemical structuring.     

Synchronization for IR‐UWB System Using a Switching Phase Detector‐Based Impulse Phase‐Locked Loop

Conventional synchronization algorithms for impulse radio require high‐speed sampling and a precise local clock. Here, a phase‐locked loop (PLL) scheme is introduced to acquire and track periodical impulses. The proposed impulse PLL (iPLL) is analyzed under an ideal Gaussian noise channel and multipath environment. The timing synchronization can be recovered directly from the locked frequency and phase. To make full use of the high harmonics of the received impulses efficiently in synchronization, the switching phase detector is applied in iPLL. It is capable of obtaining higher loop gain without a rise in timing errors. In different environments, simulations verify our analysis and show about one‐tenth of the root mean square errors of conventional impulse synchronizations. The developed iPLL prototype applied in a high‐speed ultra‐wideband transceiver shows its feasibility, low complexity, and high precision.     

Diphenylamine‐Substituted Cruciform Oligo(phenylene vinylene): Enhanced One‐ and Two‐Photon Excited Fluorescence in the Solid State

1,4‐di(4′‐N,N‐diphenylaminostyryl)benzene (DPA‐DSB) is a well known compound with a large two‐photon absorption (TPA) section and strong fluorescence in solution. However, the ease with which it crystallizes results in the formation of discontinuous crystalline phases during vacuum deposition processes, thereby greatly limiting its applicability in solid‐state devices. A cruciform dimer of DPA‐DSB, 2,5,2′,5′‐tetra(4′‐N,N‐diphenylaminostyryl)biphenyl (DPA‐TSB) is reported, wherein two DPA‐DSB molecules are linked through a central biphenyl bond. The DPA‐TSB molecules take on a cruciform configuration because of the steric crowding around the central biphenyl core, which has the effect of efficiently suppressing crystalline and intermolecular interactions. The neat DPA‐TSB solid shows strong green–blue fluorescence because of both steady‐state absorption as well as TPA. The DPA‐TSB solid exhibits a photoluminescence (PL) efficiency (η_solid) of 29 % and a solid‐state two‐photon action cross section (δη_solid) of 954 GM (1 GM = 1 × 10^–50 cm⁴ s photon^–1 molecule^–1), which is much greater than for the model compound DPA‐DSB (η_solid = 16 % and δη_solid = 150 GM, where δ is the TPA cross section and η is the fluorescence quantum yield). Based on its high PL efficiency, good film‐forming ability, and strong TPA, DPA‐TSB seems to be a good candidate for applications in solid‐state optical devices.     

Fabrication and Characterization of Two‐Photon Polymerized Features in Colloidal Crystals

The fabrication and characterization of two‐photon polymerized features written within and outside of colloidal crystals is presented. Two‐photon polymerization (TPP) response diagrams are introduced and developed to map the polymerization and damage thresholds for features written via modulated beam rastering. The use of tris[4‐(7‐benzothiazol‐2‐yl‐9,9‐diethylfluoren‐2‐yl)phenyl]amine (AF‐350) as an initiator for TPP is demonstrated for the first time and TPP response diagrams illustrate the polymerization window. These diagrams also demonstrate that the polymerization behavior within and outside of colloidal crystals is similar and electron microscopy reveals nearly identical resolution. Fluorescence confocal microscopy further enables visualization of non‐self‐supporting, three‐dimensional TPP features within self‐assembled photonic crystals. Finally, microspot spectroscopy is collected from a two‐photon feature written within a colloidal crystal and this is compared with simulation.     

Shape‐Controlled Synthesis of Pd Nanocrystals in Aqueous Solutions

This article provides an overview of recent developments regarding synthesis of Pd nanocrystals with well‐controlled shapes in aqueous solutions. In a solution‐phase synthesis, the final shape taken by a nanocrystal is determined by the twin structures of seeds and the growth rates of different crystallographic facets. Here, the maneuvering of these factors in an aqueous system to achieve shape control for Pd nanocrystals is discussed. L ‐ascorbic acid, citric acid, and poly(vinyl pyrrolidone) are tested for manipulating the reduction kinetics, with citric acid and Br^– ions used as capping agents to selectively promote the formation of {111} and {100} facets, respectively. The distribution of single‐crystal versus multiple‐twinned seeds can be further manipulated by employing or blocking oxidative etching. The shapes obtained for the Pd nanocrystals include truncated octahedron, icosahedron, octahedron, decahedron, hexagonal and triangular plates, rectangular bar, and cube. The ability to control the shape of Pd nanocrystals provides a great opportunity to systematically investigate their catalytic, electrical, and plasmonic properties.     

Two‐Photon Nanolithography Enhances the Performance of an Ionic Liquid–Polymer Composite Sensor

Continuous development of fabrication technologies, such as two‐photon polymerization (2PP), allows the exact reconstruction of specific volume shapes at micro‐ and nanometer precision. Advancements in the engineering of new materials, such as ionic liquids (ILs), are bringing superior advantages in terms of material characteristics, facilitating a combination of optical and electrical properties, as well as lithographic capabilities. In this paper, 2PP is utilized for structuring of a novel IL–polymer composite in a single‐step manufacturing process with high resolution, down to 200 nm, and high aspect ratio, up to 1:20. The composition, based on a photosensitive photoresist (e.g., IP‐L 780 or SU‐8) and the IL 1‐butyl‐3‐methylimidazolium dicyanamide, possesses a good ionic conductivity (in the range of 1–10 mS cm^?1) over a wide frequency bandwidth (1 kHz–1 MHz), an electrochemical window of 2.7 V, and a good optical transparency (transmission value of 90% for a 170 μm thick film). The fabricated structures are characterized and the phenomenon of enhanced conductivity (up to 4 S cm^?1) is explained. Two potential applications, including temperature and relative humidity sensing, are demonstrated as examples. The results suggest a new advanced approach for material structuring that can be regarded as highly most promising for a wide range of applications.     

Aggregation‐Enhanced Fluorescence and Two‐Photon Absorption in Nanoaggregates of a 9,10‐Bis[4′‐(4″‐aminostyryl)styryl]anthracene Derivative

This paper reports the design, synthesis, and theoretical modeling of two‐photon properties of a new class of chromophore that exhibits enhanced two‐photon absorption (TPA) and subsequently generated strong up‐converted emission in nanoaggregate forms. This chromophore utilizes the basic structural unit of 9,10‐bis[4′‐(4″‐aminostyryl)styryl]anthracene that exhibits large internal rotation in the monomer form in organic solvents, whereby the fluorescence is greatly reduced. In nanoaggregates formed in water, the internal rotation is considerably hindered, leading to significant increases of TPA and fluorescence quantum yield. Theoretical modeling of the conformational structure and dynamics has utilized a semiempirical pm3 formalism. The TPA cross sections of the monomer and the aggregate states have been calculated on the basis of the quadratic response theory applied to a single‐determinant self‐consistent field reference state making use of a split‐valence 6‐31G* basis set.     

Two‐Color Emitting Colloidal Nanocrystals as Single‐Particle Ratiometric Probes of Intracellular pH

Intracellular pH is a key parameter in many biological mechanisms and cell metabolism and is used to detect and monitor cancer formation and brain or heart diseases. pH‐sensing is typically performed by fluorescence microscopy using pH‐responsive dyes. Accuracy is limited by the need for quantifying the absolute emission intensity in living biological samples. An alternative with a higher sensitivity and precision uses probes with a ratiometric response arising from the different pH‐sensitivity of two emission channels of a single emitter. Current ratiometric probes are complex constructs suffering from instability and cross‐readout due to their broad emission spectra. Here, we overcome such limitations using a single‐particle ratiometric pH probe based on dot‐in‐bulk CdSe/CdS nanocrystals (NCs). These nanostructures feature two fully‐separated narrow emissions with different pH sensitivity arising from radiative recombination of core‐ and shell‐localized excitons. The core emission is nearly independent of the pH, whereas the shell luminescence increases in the 3–11 pH range, resulting in a cross‐readout‐free ratiometric response as strong as 600%. In vitro microscopy demonstrates that the ratiometric response in biologic media resembles the precalibralation curve obtained through far‐field titration experiments. The NCs show good biocompatibility, enabling us to monitor in real‐time the pH in living cells.     

Cooperative Enhancement of Two‐Photon‐Absorption‐Induced Photoluminescence from a 2D Perovskite‐Microsphere Hybrid Dielectric Structure

Two‐photon‐absorption‐induced photoluminescence (TPL) from nanostructures is generally inefficient since it is a typical third‐order nonlinear optical process. Herein, a hybrid dielectric structure composed of dielectric microspheres (approximately micrometers in diameter) covering a 2D perovskite flake is reported, which provides a straightforward strategy for enhancing the TPL emission. The microspheres in the hybrid dielectric structure not only concentrate the pumping laser but also effectively increase the detection efficiency of the emitted TPL signal. The internal quantum efficiency of the 2D perovskite is also increased in the hybrid dielectric structure due to a reduced nonradiative rate. These effects cooperatively increase the TPL emission by two orders of magnitude in the hybrid dielectric structure. Moreover, the hybrid dielectric structure is proven to be useful for TPL‐based superresolution imaging at a relatively low excitation power of 0.05 mW. This work demonstrates great promise for developing low‐cost, high‐performance nonlinear nanodevices based on hybrid dielectric structures.