Understanding the two-photon spectral properties of photoinitiators is crucial for the advancements and applications of photopolymerization (PP) techniques, specifically focusing on two-photon polymerization (TPP) for three-dimensional printing (3DP). In this work, we have investigated a specific photoinitiator designated as TECEB, a three-branched carbazole-based compound used in TPP at visible wavelengths. The lack of detailed two-photon absorption (TPA) data for TECEB is now fulfilled with this one- and two-photon absorption investigation from the experimental and theoretical points of view. Femtosecond pulses were used to acquire the TPA cross-section (sigma(2)) spectra via the Z-scan technique. Using the Gaussian software package, quantum chemical calculations (QCC) were conducted with 36 different combinations, comprising 6 chemistry models and 6 basis sets. TECEB's TPA results clearly show that this compound has a strong potential to be used as polymeric solutions in 3DP-TPP, mainly due to its TPA cross section in the spectral region between 500 nm and 550 nm, with values up to 250 GM.

A quantum-classical protocol that incorporates Jahn-Teller vibronic coupling effects and cluster analysis of molecular dynamics simulations is reported, providing a tool for simulations of absorption spectra and ultrafast nonadiabatic dynamics in large molecular photosystems undergoing aggregation in solution. Employing zinc phthalocyanine dyes as target systems, we demonstrated that the proposed protocol provided fundamental information on vibronic, electronic couplings and thermal dynamical effects that mostly contribute to the absorption spectra lineshape and the fluorescence quenching processes upon dye aggregation. Decomposing the various effects arising upon dimer formation, the structure-property relations associated with their optical responses have been deciphered at atomistic resolution.

Abstract Background: The quantitative analysis of cell surface antigens has attracted increasing attention due to the antigenic variation recognition that can facilitate diagnoses. The whole-cell-based analysis minimizes sample size, suggesting an alternative approach for detecting the variation in cell surface antigens. The optical tweezers (OT) are practical for precise single-cell manipulations, and has been widely used as a convenient tool in interdisciplinary fields. However, employing this technology as a biosensor using a "tearing" operation is still rare. Method:This paper presents a novel methodology based on the “tearing” operation of optical tweezers (OT) incorporated with the “dilution method” of antibodies to detect variations in red blood cell (RBC) surface antigens. The RBCs attach to the corresponding antibody-coated cover glass. Then, the binding firmness between an RBC and the functionalized surface is assayed by optically tearing using gradually reduced laser powers incorporated with serial antibody dilutions. Results:The experiment result shows that the higher dilution (lower antibody concentration), the lower power (lower optical force) needed to tear off the RBC binding from the functionalized surface, i.e., the antibody dilution fold is inversely proportional to the laser power. With the relative-quantitative analysis, the variation in RBC surface antigens can be intuitively estimated by comparing the maximum allowable dilution folds. The estimation gives that the antigens on the B3-type RBC are 35.7% of that on the B-type RBC, which is consistent with the literature findings using conventional biological methods. Conclusions:This study proposes a new application of the optical tweezers as a biosensor using the optically cell-tearing operation to estimate the variation in RBC surface antigens. With the proposed methodology, the detection of antigenic variation was successfully implemented without complicated optical force calculations and labored biological processing. One drop of blood from the fingertip is more than enough to obtain a satisfactory detection. It suggests an novel approach for antigenic variation analyses based on the whole-cell operation.

The quantitative analysis of cell surface antigens has attracted increasing attention due to the antigenic variation recognition that can facilitate early diagnoses. This paper presents a novel methodology based on the optical "cell-tearing" and the especially proposed "dilution regulations" to detect variations in cell surface antigens. The cell attaches to the corresponding antibody-coated slide surface. Then, the cell-binding firmness between a single cell and the functionalized surface is assayed by optically tearing using gradually reduced laser powers incorporated with serial antibody dilutions. Groups B and B3 of red blood cells (RBCs) were selected as the experiment subject. The results indicate that a higher dilution called for lower power to tear off the cell binding. According to the proposed relative-quantitative analysis theory, antigenic variation can be intuitively estimated by comparing the maximum allowable dilution folds. The estimation result shows good consistency with the finding in the literature. This study suggests a novel methodology for examining the variation in cell surface antigens, expected to be widely capable with potential sensor applications not only in biochemistry and biophysics, but also in the micro-/nano- engineering field.

Although additive manufacturing using multi-photon direct laser writing is nowadays considered as a major tool in the fabrication of future nano/micro-objects and optical components, it is currently limited by the low throughput of the writing process. To circumvent this issue, massive parallelization of the write process is a very promising avenue. However, simultaneous writing of structures in close spatial proximity generates fabrication artefacts, collectively referred to as “proximity effects”, which strongly limit the accessible structure resolution. In this work, we systematically investigate the experimental parameters that influence these effects using specifically designed N×N spot diffractive optical elements. Through computer simulations, we show that these effects can be modeled remarkably successfully simply by taking Point Spread Function overlap and diffusion processes into account. We illustrate the usefulness of the concept by designing a parallel write approach giving access to periodic structures with short inter-object distances while very largely overcoming proximity effects.

We elucidate the crucial role of the cetyl trimethylammonium bromide (CTAB) surfactant in the anisotropic growth mechanism of gold nano-bipyramids, nano-objects with remarkable optical properties and high tunability. Atomistic molecular dynamics simulations predict different surface coverages of the CTAB (positively charged) heads and their (bromide) counterions as function of the gold exposed surfaces. High concentration of CTAB surfactant promotes formation of gold nanograins in solution that work as precursors for the smooth anisotropic growth of more elongated nano-bipyramidal objects. Nanobipyramids feature higher index facets with respect to nanorods, allowing higher CTAB coverages that stabilize their formation and leading to narrower inter-micelles channels that smooth down their anisotropic growth. Absorption spectroscopy and scanning electron microscopy confirmed the formation of nanograins and demonstrated the importance of surfactant concentration on driving the growth towards nano-bipyramids rather than nanorods. The outcome explains the formation of the monodisperse bipyramidal nano-objects, the origin of their controlled shapes and sizes along with their remarkable stability.

Two-photon lithography is a potential route to produce high-resolution 3D ceramics. However, the large shrinkage due to the elimination of an important organic counterpart of the printed material during debinding/sintering remains a lock to further development of this technology. To limit this phenomenon, an original approach based on a composite resin incorporating 45 wt% ultrasmall (5 nm) zirconia stabilized nanoparticles into the zirconium acrylate precursor is proposed to process 3D zirconia microlattices and nanostructured optical surfaces. Interestingly, the nanoparticles are used both as seeds allowing control of the crystallographic phase formed during the calcination process and as structural stabilizing agent preventing important shrinkage of the printed ceramic. After 3D photolithography and pyrolysis, the weight and volume loss of the microstructures are drastically reduced as compared to similar systems processed with the reference resin without nanoparticles, and stable 3D microstructures of cubic zirconia are obtained with high spatial resolution. In the case of a patterned surface, the refractive index of 2.1 leads to a diffraction efficiency large enough to obtain microfocusing with linewidths of 0.1 µm, and the demonstration of a microlens array with a period as small as 0.8 µm.

Recent developments in 3D micromixers promise to accomplish satisfactory mixing for applications in microchemical process, biomicrofluidic analyses, m-TAS, etc. However, the complicated 3D architecture employed to ensure thorough mixing might reduce fluid circulation due to the space for fluid flowing becomes narrow. In this study, we developed a single 3D multi-manifold micromixer composed of a doubleArchimedes-screw for rapidly mixing within a short distance. Archimedes screw is well known for its fluid circulation under stable flow. The mixing strategy is based on split-and-recombination for increasing the interfacial area, and the 3D flow rotation for generating omnidirectional momentum-changes during the fluid propagation. With the spiral conformation, the Archimedes-screw micromixer (ASM) can substantially enhance fluid circulation. Numerical simulations used to help to design the optimal screw-turn show that satisfactory mixing efficiency can achieve at the distance of approximately twice the microchannel width. The ASM characteristics, such as pressure loss, ratio of cross-sections, and Reynolds number (Re) were evaluated. Experimental demonstration by an ASM fabricated by two-photon stereolithography showed mixing result in good agreement with the simulation findings. (c) 2021 Taiwan Institute of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

Two-photon polymerization is a photochemical process usually initiated by tightly focusing an ultrafast laser pulse into a volume of photosensitive photoresists with a high-numerical-aperture objective. Scanning a write "voxel" in 3D enables near free-form fabrication, but at a limited speed which is a critical factor for industrial purposes, because generally only a single writing-beam is used. Several strategies have been implemented to improve the fabrication speed, one such strategy is massive parallelization which is the approach used in our PHENOmenon H2020 European project. Massive parallelization can be realized by beam splitting diffractive optical elements which allow simultaneous fabrication with thousands of beams, decreasing the overall fabrication time. A major unexpected obstacle is encountered in massively parallelized fabrication: using several spots simultaneously to polymerize, local changes in the 2PP threshold have been observed. We linked this to the proximity effect. The aim of this study is to understand the proximity effect in parallel microfabrication using simulation to predict its behaviour and different systematic experiments to reduce the proximity effect such as changing photoresist, using thinner photoresist layers to increase oxygen penetration or using higher Numerical Aperture Objectives.

High resolution fabrication using two-photon lithography is extensively studied for a large range of materials, from polymer to inorganics. Hybrid materials including a sol–gel step have been developed since two decades to increase mechanical or optical properties in particular on silicon-based materials. Among the metal oxide, few studies have been dedicated to titanium and, because of the high reactivity of titanium precursors, obtaining a resin with a high part of titanium is challenging. Indeed, resins for two-photon lithography have to be stable for the processing time and titanium precursors are more difficult to operate due to their higher reactivity and often require extreme working conditions in order to control the chemical processes. Here, we propose a method, working at ambient conditions, to print submicronic structures of organic–inorganic hybrids with a large proportion of titanium and ceramics using high resolution two-photon process. The material obtained and its evolution during the pyrolysis at 600 and 1000 °C are characterized. We show that TiO 2 /SiO 2 -based microceramics can be obtained after the pyrolysis of the microstructures. The respective roles of the two chemical reactions involved in this lithography process, sol–gel condensation and radical photopolymerization, are highlighted.

The use of two-photon absorption (TPA) for polymerization, also known as 3D Lithography, Direct Laser Writing, or High-Precision 3D Printing is gaining increasing attraction in industrial fabrication of micro- and nanostructures. Mainly due to its vast freedom in design and high-resolution capabilities, TPA enables the fabrication of designs which are not feasible or far too complicated to be achieved with conventional fabrication methods. TPA is a scanning technology and fabrication in 3D requires axial overwritings. High industrial throughput fabrication can be achieved by intelligent fabrication strategies combined with an excellent material basis. Further boosting the throughput can be achieved by multispot exposure strategies. In this paper, massive parallelization is demonstrated which was realized by using a beam splitting diffractive optical element (DOE). Simultaneous fabrication using commercially available acrylate-based hybrid resin with 121 parallel focal spots arranged as 11 x 11 array is reported. Structures fabricated by a single laser beam and by 121 parallel beams are compared to each other with regard to shape and polymerization threshold. It was found that polymerization is strongly increased when parallel beams are used, especially for the central beams. As a result, polymerization threshold is lower in the center of the 11 x 11 array compared to the edges of the array. Furthermore, structures at the center of the 11 x 11 array are bigger compared to structures at the edges of the array when assigning equal intensity to all diffracted beams. These results are attributed to diffusion of photo initiators, quenchers, and radicals.

The quantitative analysis of surface antigens on cells, especially red blood cells (RBCs), has attracted increasing attention due to the recognition of antigenic changes that can facilitate early diagnoses. This paper presents an alternative methodology developed using the optical cell-detachment technique to evaluate antibody-antigen interactions and quantitatively analyze the RBC surface antigen expression. RBC subtyping was used to verify the proposed detection principle based on a comparison of the bonding strengths between individual RBCs and antibody coatings. The bonding strengths were measured with serial antibody dilutions with gradually decreasing laser powers, for which a single cell was optically detached from the corresponding antibody-coated surface. With the quantitatively analysis, the proposed alternative methodology was verified as a highly sensitive technique for detecting antigen expression on the RBC surface.

Diffractive approaches are needed when refractive microlenses reach their focusing limit at the micron-scale in visible light. Previously, we have reported on micron-sized optical lenses based on the diffraction of metallic nanowires. Here, we extend our study to lenses based on pairs of subwavelength dielectric scatterers. Using simulations by two-dimensional finite element method, we demonstrate that focusing holds for pair spacings as small as the wavelength-size. For pairs with distances between inner walls larger than about 1.2λ, the scattered waves generate a quadratic phase modulation on the total propagating field leading to a diffraction-limited focusing i.e. an effective optical lens effect with high numerical aperture. In addition, they have low sidelobe intensities, long depths of focus, and they have a low sensitivity with polarization. For pairs with inner wall distances smaller than about 1.2λ, the focusing phase modulation is accumulated during the propagation through the dielectric pair structure. In this work, we report only on the experimental demonstration for the case of larger wall separation to emphasize on the scattered wave effect on micro-focusing. A pair of parallel polymer lines (cylindrical lens), and a grid of polymer lines (square microlens array) with 2 μm-spacing were fabricated by two-photon induced polymerization. Their focal lengths are comparable to their separating distances, their spot-sizes are 0.37 μm and 0.28 μm at wavelength 530 nm, and their focusing efficiencies are 70% and 60%, respectively.

Epicocconone 1 is a natural chromophore isolated from the fungus Epicoccum nigrum that has shown applications in proteomics and fluorescent microscopy thanks to its unique pro-fluorescence properties. The modification of the skeleton of the natural product by replacing the triene side chain by a fluorenyl scaffold can noticeably increase the fluorophore's absorption coefficient. The synthesis of the analogues of the natural product has been made possible by the use of a palladium catalyzed carbonylation reaction, allowing the construction of the β-keto-dioxinone key intermediate. Two-photon absorption cross section measurements of the fluorenyl epicocconone analogues show a structure-dependency with values ranging from 60 GM to 280 GM and live cell imaging show intense staining of intracellular vesicles-like structures around the nucleus.

A large series of structurally related two-photon photosensitizers with heavy atom substitution were synthesized and evaluated through a combined spectroscopic (steady-state and time resolved), photophysical and computational study. Our aim was to identify some relevant parameters related to their excited state dynamics including photo-induced singlet oxygen generation. Although these dynamics result from the interplay of many factors, we show that the triplet excited state generation kinetics can generally be correlated with the calculated values of both the spin-orbit coupling and the energy gap between S1 and T1 states, which themselves mostly depend on the positioning of the heavy atoms along the π-conjugated structure rather than their number.

One of the challenges of photodynamic therapy is to increase the penetration depth of light irradiation in the tumor tissues. Although two-photon excitation strategies have been developed, the two-photon absorption cross sections of clinically used photosensitizers are generally low (below 300 GM). Besides, photosensitizers with high cross section values are often non-water-soluble. In this research work, a whole family of photosensitizer-polymer conjugates was synthesized via the covalent binding of a photosensitizer with a relatively high cross section along a biocompatible copolymer chain. The resulting photosensitizer-polymer conjugates were water-soluble and could be imaged in cellulo by two-photon microscopy thanks to their high two-photon absorption cross sections (up to 2600 GM in water, in the NIR range). In order to explore the structure/photodynamic activity relationship of such macromolecular photosensitizers, the influence of the polymer size, photosensitizer density, and presence of charges along the polymer backbone was investigated (neutral, anionic, cationic, and zwitterionic conjugates were compared). The macromolecular photosensitizers were not cytotoxic in the absence of light irradiation. Their kinetics of cellular uptake in the B16-F10 melanoma cell line were followed by flow cytometry over 24 h. The efficiency of cell death upon photoactivation was found to be highly correlated to the cellular uptake in turn correlated to the global charge of the macromolecular photosensitizer which appeared as the determining structural parameter.

Two-photon excitation of photobases is used to induce pH changes and control the condensation step of the sol–gel process at the focal point of a laser beam in a confocal configuration.

This study fabricated periodic 3D nanostructures by using a periodic voxel array for light extraction.Two-photon polymerization (TPP) is a well-known technology used for generating complex 3D micro-and nanostructures with sub-100-nm resolution.Because of the periodic voxel-by-voxel array fabrication process, the fabrication of holographic two-photon polymerization (HTPP) can be faster than that of traditional TPP.HTPP promises a flexible technique to fabricate an arbitrary periodic 3D structured optical surface.In this research, we used a 25 × 25 beam-splitting holographic mask to generate a periodic focus in a photocured material and to systematically fabricate complex 3D nanostructures at rates of mm² per minute.The optical responses of nanostructures with a 1 μm period fabricated in a polymer were characterized using 3D optical microscopy.The experimental results of the light distribution demonstrated favorable agreement with those of the simulation.In the light-extracting experiment, the light extraction capability of the gold-layer nanostructures was 3.6 times higher than that of the glass without nanostructures.

Dyes emitting in the solid state in the red or near-infrared range are much sought after for application in bioimaging especially if the long emission wavelength can be combined with two-photon excitation to provide unique contrast and penetration depth.