Fabrication of multilevel metalenses using multiphoton lithography: from design to evaluation.

We present a procedure for the design of multilevel metalenses and their fabrication with multiphoton-based direct laser writing. This work pushes this fast and versatile fabrication technique to its limits in terms of achievable feature size dimensions for the creation of compact high-numerical aperture metalenses on flat substrates and optical fiber tips. We demonstrate the design of metalenses with various numerical apertures up to 0.96, and optimize the fabrication process towards nanostructure shape reproducibility. We perform optical characterization of the metalenses towards spot size, focusing efficiency, and optical functionality with a fiber beam collimation design, and compare their performance with refractive and diffractive counterparts fabricated with the same technology.

Mass-manufacturable scintillation-based optical fiber dosimeters for brachytherapy.

Scintillation-based fiber dosimeters are a powerful tool for minimally invasive localized real-time monitoring of the dose rate during Low Dose Rate (LDR) and High Dose Rate (HDR) brachytherapy (BT). This paper presents the design, fabrication, and characterization of such dosimeters, consisting of scintillating sensor tips attached to polymer optical fiber (POF). The sensor tips consist of inorganic scintillators, i.e. Gd2O2S:Tb for LDR-BT, and Y2O3:Eu+4YVO4:Eu for HDR-BT, dispersed in a polymer host. The shape and size of the tips are optimized using non-sequential ray tracing simulations towards maximizing the collection and coupling of the scintillation signal into the POF. They are then manufactured by means of a custom moulding process implemented on a commercial hot embossing machine, paving the way towards series production. Dosimetry experiments in water phantoms show that both the HDR-BT and LDR-BT sensors feature good consistency in the magnitude of the average photon count rate and that the photon count rate signal is not significantly affected by variations in sensor tip composition and geometry. Whilst individual calibration remains necessary, the proposed dosimeters show great potential for in-vivo dosimetry for brachytherapy.

Design and demonstration of a six-channel multiresolution imaging system.

Multichannel imaging systems consist of multiple channels that have different imaging characteristics (fields-of-view and angular resolutions). We design and demonstrate a six-channel multiresolution imaging system that can achieve a relative magnification ratio of up to 10 times between the channels and at the same time result in different depths-of-field. The imaging system consists of two double-sided lens arrays made of PMMA material, a baffle to eliminate possible cross talk between neighboring channels, and a Sony full-frame image sensor. The imaging system was tested by capturing images of stationary and moving objects. The acquired images exhibit different resolutions, fields-of-view, and levels of blur proving our concept.

On the potential use of two-photon polymerization to 3D print chromatographic packed bed supports.

The continuous quest for chromatographic supports offering kinetic performance properties superior to that of the packed bed of spheres has pushed the field to consider alternative formats such as for example monolithic and pillar array columns. This quest seems bound to culminate in the use of 3D printing technology, as this intrinsically offers the possibility to produce supports with a perfect uniformity and with a size and shape that is fully optimized for the chromatographic separation process. However, to be competitive with the current state-of-the-art, structures with sub-micron feature sizes are required. The present contribution therefore investigates the use of the 3D printing technology with the highest possible resolution available today, i.e., two-photon polymerization (2PP). It is shown that 2PP printing is capable of achieving the required ≤ 1 µm printing resolution. Depending on the laser scan speed, the lower limit through-pore size for a tetrahedral skeleton monolith with a theoretical 80% external porosity was found to be at 800 nm, when printing at a scan speed of 50 mm/s with a laser power of 10%. For a scan speed of 10 mm/s, the minimal through-pore size dropped to 500 nm. However, this very high resolution comes at the cost of excessively long printing times. The total printing time for a column volume equivalent to that of a typical nano-LC column (75 µm i.d. cylindrical tube with length L = 15 cm) has been determined to correspond to 330 and 470 h for the 50 mm/s and the 10 mm/s scan speed respectively. Other issues remaining to be solved are the need to clad the printed skeleton with a suitable mesoporous layer for chromatographic retention and the need to add a top-wall to the printed channels after the removal of the non-polymerized resin. It is therefore concluded that 2PP printing is not ready yet to replace the existing column fabrication methods.

Increasing the Microfabrication Performance of Synthetic Hydrogel Precursors through Molecular Design.

Implementation of hydrogel precursors in two-photon polymerization (2PP) technology provides promising opportunities in the tissue engineering field thanks to their soft characteristics and similarity to extracellular matrix. Most of the hydrogels, however, are prone to post-fabrication deformations, leading to a mismatch between the computer-aided design and the printed structure. In the present work, we have developed novel synthetic hydrogel precursors to overcome the limitations associated with 2PP processing of conventional hydrogel precursors such as post-processing deformations and a narrow processing window. The precursors are based on a poly(ethylene glycol) backbone containing urethane linkers and are, on average, functionalized with six acrylate terminal groups (three on each terminal group). As a benchmark material, we exploited a precursor with an identical backbone and urethane linkers, albeit functionalized with two acrylate groups, that were reported as state-of-the-art. An in-depth characterization of the hexafunctional precursors revealed a reduced swelling ratio (<0.7) and higher stiffness (>36 MPa Young's modulus) compared to their difunctional analogs. The superior physical properties of the newly developed hydrogels lead to 2PP-based fabrication of stable microstructures with excellent shape fidelity at laser scanning speeds up to at least 90 mm s-1, in contrast with the distorted structures of conventional difunctional precursors. The hydrogel films and microscaffolds revealed a good cell interactivity after functionalization of their surface with a gelatin methacrylamide-based coating. The proposed synthesis strategy provides a one-pot and scalable synthesis of hydrogel building blocks that can overcome the current limitations associated with 2PP fabrication of hydrogel microstructures.

3D direct laser writing of microstructured optical fiber tapers on single-mode fibers for mode-field conversion.

We present a design and fabrication approach for 3D printed polymer microstructured optical fiber tapers on standard single-mode glass fibers for efficient and compact mode-field conversion. This paves the way towards complex functionalized fiber tips for various applications, like sensors and beam shaping components, currently limited by the mode-field size and distribution of standard optical fibers. In this paper, we demonstrate the potential of mode-field converting tapers for relaxing the misalignment tolerance in fiber-to-fiber connections and maximizing the coupling efficiency in fiber-to-chip connections. We demonstrate a mode-field diameter expansion ratio of 1.7 and reduction ratio of 3 and show that our microstructured tapers achieve a comparable performance in coupling efficiency as their step-index counterparts, while providing greater robustness.

Thiol-norbornene gelatin hydrogels: influence of thiolated crosslinker on network properties and high definition 3D printing.

Photocrosslinkable gelatin hydrogels are excellent bioinks or biomaterial ink components to serve biofabrication applications. Especially the widely investigated gelatin-methacroyl (gel-MA) hydrogels hold an impressive track record. However, over the past decade, increasing attention is being paid to thiol-ene photo-click chemistry to obtain hydrogel networks benefitting from a faster reactivity (i.e. seconds vs minutes) along with superior biocompatibility and processability. In order to exploit this photo-click chemistry, often an ene-functionality (e.g. norbornene) is introduced onto gelatin followed by crosslinking in the presence of a multifunctional thiol (e.g. dithiothreitol). To date, very limited research has been performed on the influence of the applied thiolated crosslinker on the final hydrogel properties. Therefore, the present work assesses the influence of different thiolated crosslinkers on the crosslinking kinetics, mechanical properties and biological performance of the hydrogels upon encapsulation of primary adipose tissue-derived stem cells which indicated a cell viability exceeding 70%. Furthermore, the different formulations were processed using two-photon polymerization which indicated, in addition to differences in processing window and swelling ratio, a previously unreported phenomenon. At high intensities (i.e. ⩾150 mW), the laser results in cleavage of the gelatin backbone even in the absence of distinct photo-cleavable functionalities. This can have potential to introduce channels or softer regions in gels to result in zones characterized by different degradation speeds or the formation of blood vessels. Consequently, the present study can be used to provide guidance towards tailoring the thiol-ene system towards the desired applications.

Measurement of the soliton number in guiding media through continuum generation.

No general approach is available yet to measure directly the ratio between chromatic dispersion and the nonlinear coefficient, and hence the soliton number for a given optical pulse, in an arbitrary guiding medium. Here we solve this problem using continuum generation. We experimentally demonstrate our method in polarization-maintaining and single-mode fibers with positive and negative chromatic dispersion. Our technique also offers new opportunities to determine the chromatic dispersion of guiding media over a broad spectral range while pumping at a fixed wavelength.

Designer Descemet Membranes Containing PDLLA and Functionalized Gelatins as Corneal Endothelial Scaffold.

Corneal blindness is the fourth leading cause of visual impairment. Of specific interest is blindness due to a dysfunctional corneal endothelium which can only be treated by transplanting healthy tissue from a deceased donor. Unfortunately, corneal supply does not meet the demand with only one donor for every 70 patients. Therefore, there is a huge interest in tissue engineering of grafts consisting of an ultra-thin scaffold seeded with cultured endothelial cells. The present research describes the fabrication of such artificial Descemet membranes based on the combination of a biodegradable amorphous polyester (poly (d,l-lactic acid)) and crosslinkable gelatins. Four different crosslinkable gelatin derivatives are compared in terms of processing, membrane quality, and function, as well as biological performance in the presence of corneal endothelial cells. The membranes are fabricated through multi-step spincoating, including a sacrificial layer to allow for straightforward membrane detachment after production. As a consequence, ultrathin (<1 µm), highly transparent (>90%), semi-permeable membranes could be obtained with high biological potential. The membranes supported the characteristic morphology and correct phenotype of corneal endothelial cells while exhibiting similar proliferation rates as the positive control. As a consequence, the proposed membranes prove to be a promising synthetic alternative to donor tissue.

SERS using two-photon polymerized nanostructures for mycotoxin detection.

Improved chemical- and bio-sensing with Surface Enhanced Raman Spectroscopy (SERS) requires nanostuctures that can be flexibly designed and fabricated with different physical and optical properties. Here, we present nano-pillar arrays ranging from 200 nm to 600 nm as SERS substrates for mycotoxin detection that are fabricated by means of two-photon polymerization. We built a nominal shape and a voxel-based model for simulating the enhancement of the electric field of the nano-pillar arrays using the Finite-Difference Time-Domain (FDTD) method. A new model was built based on the Atomic Force Microscopy (AFM) data obtained from the fabricated nanostructures and introduced into a FDTD model. We demonstrated the enhancement behavior by measuring the Raman spectrum of Rhodamine B solutions. Both the simulations and experimental results suggest that the 200 nm nano-pillar array has the highest Enhancement Factor (EF). Besides, we determined the limit of detection of the 200 nm pillar array by performing Raman measurements on Rhodamine B solutions with different concentrations. The detection limit of our 200 nm nano-pillar array is 0.55 µM. Finally we discriminated 1 ppm deoxynivalenol and 1.25 ppm fumonisin b1 in acetonitrile solutions by our SERS substrate in combination with principal component analysis. This versatile approach for SERS substrates fabrication gives new opportunities for material characterization in chemical and biological applications.

(Photo-)crosslinkable gelatin derivatives for biofabrication applications.

Over the recent decades gelatin has proven to be very suitable as an extracellular matrix mimic for biofabrication and tissue engineering applications. However, gelatin is prone to dissolution at typical cell culture conditions and is therefore often chemically modified to introduce (photo-)crosslinkable functionalities. These modifications allow to tune the material properties of gelatin, making it suitable for a wide range of biofabrication techniques both as a bioink and as a biomaterial ink (component). The present review provides a non-exhaustive overview of the different reported gelatin modification strategies to yield crosslinkable materials that can be used to form hydrogels suitable for biofabrication applications. The different crosslinking chemistries are discussed and classified according to their mechanism including chain-growth and step-growth polymerization. The step-growth polymerization mechanisms are further classified based on the specific chemistry including different (photo-)click chemistries and reversible systems. The benefits and drawbacks of each chemistry are also briefly discussed. Furthermore, focus is placed on different biofabrication strategies using either inkjet, deposition or light-based additive manufacturing techniques, and the applications of the obtained 3D constructs. STATEMENT OF SIGNIFICANCE: Gelatin and more specifically gelatin-methacryloyl has emerged to become one of the gold standard materials as an extracellular matrix mimic in the field of biofabrication. However, also other modification strategies have been elaborated to take advantage of a plethora of crosslinking chemistries. Therefore, a review paper focusing on the different modification strategies and processing of gelatin is presented. Particular attention is paid to the underlying chemistry along with the benefits and drawbacks of each type of crosslinking chemistry. The different strategies were classified based on their basic crosslinking mechanism including chain- or step-growth polymerization. Within the step-growth classification, a further distinction is made between click chemistries as well as other strategies. The influence of these modifications on the physical gelation and processing conditions including mechanical properties is presented. Additionally, substantial attention is put to the applied photoinitiators and the different biofabrication technologies including inkjet, deposition or light-based technologies.

Technological advancements for the development of stem cell-based models for hepatotoxicity testing.

Stem cells are characterized by their self-renewal capacity and their ability to differentiate into multiple cell types of the human body. Using directed differentiation strategies, stem cells can now be converted into hepatocyte-like cells (HLCs) and therefore, represent a unique cell source for toxicological applications in vitro. However, the acquired hepatic functionality of stem cell-derived HLCs is still significantly inferior to primary human hepatocytes. One of the main reasons for this is that most in vitro models use traditional two-dimensional (2D) setups where the flat substrata cannot properly mimic the physiology of the human liver. Therefore, 2D-setups are progressively being replaced by more advanced culture systems, which attempt to replicate the natural liver microenvironment, in which stem cells can better differentiate towards HLCs. This review highlights the most recent cell culture systems, including scaffold-free and scaffold-based three-dimensional (3D) technologies and microfluidics that can be employed for culture and hepatic differentiation of stem cells intended for hepatotoxicity testing. These methodologies have shown to improve in vitro liver cell functionality according to the in vivo liver physiology and allow to establish stem cell-based hepatic in vitro platforms for the accurate evaluation of xenobiotics.

Graphene's nonlinear-optical physics revealed through exponentially growing self-phase modulation.

Graphene is considered a record-performance nonlinear-optical material on the basis of numerous experiments. The observed strong nonlinear response ascribed to the refractive part of graphene's electronic third-order susceptibility χ(3) cannot, however, be explained using the relatively modest χ(3) value theoretically predicted for the 2D material. Here we solve this long-standing paradox and demonstrate that, rather than χ(3)-based refraction, a complex phenomenon which we call saturable photoexcited-carrier refraction is at the heart of nonlinear-optical interactions in graphene such as self-phase modulation. Saturable photoexcited-carrier refraction is found to enable self-phase modulation of picosecond optical pulses with exponential-like bandwidth growth along graphene-covered waveguides. Our theory allows explanation of these extraordinary experimental results both qualitatively and quantitatively. It also supports the graphene nonlinearities measured in previous self-phase modulation and self-(de)focusing (Z-scan) experiments. This work signifies a paradigm shift in the understanding of 2D-material nonlinearities and finally enables their full exploitation in next-generation nonlinear-optical devices.

Highly Reactive Thiol-Norbornene Photo-Click Hydrogels: Toward Improved Processability.

In the present work, gelatin type B is modified with highly reactive norbornene functionalities (Gel-NB) following a one-pot synthesis approach to enable subsequent thiol-ene photo-click crosslinking. The modification strategy displays close control over the amount of introduced functionalities. Additionally, Gel-NB exhibits considerably improved processing capabilities in terms of two-photon polymerization when benchmarked to earlier-reported crosslinkable gelatin derivatives (e.g., gelatin-methacrylamide (Gel-MOD) and gelatin-methacrylamide-aminoethylmethacrylate (Gel-MOD-AEMA)). The improvement is especially apparent in terms of minimally required laser power (20 mW vs ≥60 mW (Gel-MOD) vs ≥40 mW (Gel-MOD-AEMA) at 100 mm s-1 scan speed) and processable concentration range (≥5 w/v% vs ≥10 w/v% (Gel-MOD/Gel-MOD-AEMA)). Furthermore, the proposed functionalization scheme maintains the excellent biocompatibility and cell interactivity of gelatin. Additionally, the norbornene functionalities have potential for straightforward postprocessing "thiol-ene" surface grafting of active molecules. As a consequence, a very promising material toward tissue engineering applications and more specifically, biofabrication, is presented.

Cross-Linkable Gelatins with Superior Mechanical Properties Through Carboxylic Acid Modification: Increasing the Two-Photon Polymerization Potential.

The present work reports on the development of photo-cross-linkable gelatins sufficiently versatile to overcome current biopolymer two-photon polymerization (2PP) processing limitations. To this end, both the primary amines as well as the carboxylic acids of gelatin type B were functionalized with photo-cross-linkable moieties (up to 1 mmol/g) resulting in superior and tunable mechanical properties (G' from 5000 to 147000 Pa) enabling efficient 2PP processing. The materials were characterized in depth prior to and after photoinduced cross-linking using fully functionalized gelatin-methacrylamide (gel-MOD) as a benchmark to assess the effect of functionalization on the protein properties, cross-linking efficiency, and mechanical properties. In addition, preliminary experiments on hydrogel films indicated excellent in vitro biocompatibility (close to 100% viability) both in the presence of MC3T3 preosteoblasts and L929 fibroblasts. Moreover, 2PP processing of the novel derivative was superior in terms of applied laser power (≥40 vs ≥60 mW for gel-MOD at 100 mm/s) as well as post-production swelling (0-20% vs 75-100% for gel-MOD) compared to those of gel-MOD. The reported novel gelatin derivative (gel-MOD-AEMA) proves to be extremely suitable for direct laser writing as both superior mimicry of the applied computer-aided design (CAD) was obtained while maintaining the desired cellular interactivity of the biopolymer. It can be anticipated that the present work will also be applicable to alternative biopolymers mimicking the extracellular environment such as collagen, elastin, and glycosaminoglycans, thereby expanding current material-related processing limitations in the tissue engineering field.

Indirect Rapid Prototyping: Opening Up Unprecedented Opportunities in Scaffold Design and Applications.

Over the past decades, solid freeform fabrication (SFF) has emerged as the main technology for the production of scaffolds for tissue engineering applications as a result of the architectural versatility. However, certain limitations have also arisen, primarily associated with the available, rather limited range of materials suitable for processing. To overcome these limitations, several research groups have been exploring novel methodologies through which a construct, generated via SFF, is applied as a sacrificial mould for production of the final construct. The technique combines the benefits of SFF techniques in terms of controlled, patient-specific design with a large freedom in material selection associated with conventional scaffold production techniques. Consequently, well-defined 3D scaffolds can be generated in a straightforward manner from previously difficult to print and even "unprintable" materials due to thermomechanical properties that do not match the often strict temperature and pressure requirements for direct rapid prototyping. These include several biomaterials, thermally degradable materials, ceramics and composites. Since it can be combined with conventional pore forming techniques, indirect rapid prototyping (iRP) enables the creation of a hierarchical porosity in the final scaffold with micropores inside the struts. Consequently, scaffolds and implants for applications in both soft and hard tissue regeneration have been reported. In this review, an overview of different iRP strategies and materials are presented from the first reports of the approach at the turn of the century until now.

Mass-manufacturable polymer microfluidic device for dual fiber optical trapping.

We present a microfluidic chip in Polymethyl methacrylate (PMMA) for optical trapping of particles in an 80µm wide microchannel using two counterpropagating single-mode beams. The trapping fibers are separated from the sample fluid by 70µm thick polymer walls. We calculate the optical forces that act on particles flowing in the microchannel using wave optics in combination with non-sequential ray-tracing and further mathematical processing. Our results are compared with a theoretical model and the Mie theory. We use a novel fabrication process that consists of a premilling step and ultraprecision diamond tooling for the manufacturing of the molds and double-sided hot embossing for replication, resulting in a robust microfluidic chip for optical trapping. In a proof-of-concept demonstration, we show the trapping capabilities of the hot embossed chip by trapping spherical beads with a diameter of 6µm, 8µm and 10µm and use the power spectrum analysis of the trapped particle displacements to characterize the trap strength.

Laser ablation- and plasma etching-based patterning of graphene on silicon-on-insulator waveguides.

We present a new approach to remove monolayer graphene transferred on top of a silicon-on-insulator (SOI) photonic integrated chip. Femtosecond laser ablation is used for the first time to remove graphene from SOI waveguides, whereas oxygen plasma etching through a metal mask is employed to peel off graphene from the grating couplers attached to the waveguides. We show by means of Raman spectroscopy and atomic force microscopy that the removal of graphene is successful with minimal damage to the underlying SOI waveguides. Finally, we employ both removal techniques to measure the contribution of graphene to the loss of grating-coupled graphene-covered SOI waveguides using the cut-back method.

Indirect additive manufacturing as an elegant tool for the production of self-supporting low density gelatin scaffolds.

The present work describes for the first time the production of self-supporting low gelatin density (<10 w/v%) porous scaffolds using methacrylamide-modified gelatin as an extracellular matrix mimicking component. As porous scaffolds starting from low gelatin concentrations cannot be realized with the conventional additive manufacturing techniques in the abscence of additives, we applied an indirect fused deposition modelling approach. To realize this, we have printed a sacrificial polyester scaffold which supported the hydrogel material during UV crosslinking, thereby preventing hydrogel structure collapse. After complete curing, the polyester scaffold was selectively dissolved leaving behind a porous, interconnective low density gelatin scaffold. Scaffold structural analysis indicated the success of the selected indirect additive manufacturing approach. Physico-chemical testing revealed scaffold properties (mechanical, degradation, swelling) to depend on the applied gelatin concentration and methacrylamide content. Preliminary biocompatibility studies revealed the cell-interactive and biocompatible properties of the materials developed.

Two-channel multiresolution refocusing imaging system using a tunable liquid lens.

Multichannel imaging systems currently feature refocusing capabilities only in bulky and expensive designs. Mechanical movements of the components cannot be integrated in miniaturized designs, preventing classical refocusing mechanisms. To overcome this limitation we developed, as a proof-of-concept (POC) demonstration, a compact low-cost two-channel refocusing imaging system based on a voltage-tunable liquid lens. In addition, the design can be realized with wafer-level manufacturing techniques. One channel of the imaging system enables a wide field of view (FOV) of a scene (2×40°) but with a limited angular resolution (0.078°), while the other channel gives a high angular resolution (0.0098°) image of a small region of interest but with a much narrower FOV (2×7.57°). It is this high-resolution channel that contains the tunable lens and therefore the refocusing capability. A POC demonstration of the proposed two-channel system was built and its performances were measured. Both imaging channels show good overall diffraction-limited image quality.