Quantitative phase imaging verification in large field-of-view lensless holographic microscopy via two-photon 3D printing.

Large field-of-view (FOV) microscopic imaging (over 100 mm2) with high lateral resolution (1-2 µm) plays a pivotal role in biomedicine and biophotonics, especially within the label-free regime. Lensless digital holographic microscopy (LDHM) is promising in this context but ensuring accurate quantitative phase imaging (QPI) in large FOV LDHM is challenging. While phantoms, 3D printed by two-photon polymerization (TPP), have facilitated testing small FOV lens-based QPI systems, an equivalent evaluation for lensless techniques remains elusive, compounded by issues such as twin-image and beam distortions, particularly towards the detector's edges. Here, we propose an application of TPP over large area to examine phase consistency in LDHM. Our research involves fabricating widefield phase test targets with galvo and piezo scanning, scrutinizing them under single-shot twin-image corrupted conditions and multi-frame iterative twin-image minimization scenarios. By measuring the structures near the detector's edges, we verified LDHM phase imaging errors across the entire FOV, with less than 12% phase value difference between areas. Our findings indicate that TPP, followed by LDHM and Linnik interferometry cross-verification, requires new design considerations for precise large-area photonic manufacturing. This research paves the way for quantitative benchmarking of large FOV lensless phase imaging, enhancing understanding and further development of LDHM technique.

Perspective on Water-Soluble Two-Photon Initiator for Two-Photon Polymerization.

Two-photon polymerization (TPP) as an unparalleled technology empowers the rapid prototyping of customized three-dimensional (3D) micro/nanostructures, garnering noticeable interest in tissue engineering, drug delivery, and regenerative medicine. These applications have a high requirement on the biocompatibility and integrity of 3D structures. Therefore, it is important to develop two-photon initiator with good water-solubility, initiation efficiency, and biocompatibility. Here, we share our insights into the development of a water-soluble two-photon initiator (WTPI) and applications from the material and manufacturing perspective. We highlight the nonlinear optical properties and the synthesis of WTPI through three pathways. Then we further demonstrate the applications of the TPP technique in the aqueous phase in the fields of tissue engineering, 4D printing, and ceramic manufacturing. Finally, a general conclusion and outlook are provided for the future development and application of WTPI.

2PP-Hydrogel Covered Electrodes to Compensate for Media Effects in the Determination of Biomass in a Capillary Wave Micro Bioreactor.

Microbioreactors increase information output in biopharmaceutical screening applications because they can be operated in parallel without consuming large quantities of the pharmaceutical formulations being tested. A capillary wave microbioreactor (cwMBR) has recently been reported, allowing cost-efficient parallelization in an array that can be activated for mixing as a whole. Although impedance spectroscopy can directly distinguish between dead and viable cells, the monitoring of cells in suspension within bioreactors is challenging because the signal is influenced by the potentially varying properties of the culture medium. In order to address this challenge, an impedance sensor consisting of two sets of microelectrodes in a cwMBR is presented. Only one set of electrodes was covered by a two-photon cross-linked hydrogel to become insensitive to the influence of cells while remaining sensitive to the culture medium. With this impedance sensor, the biomass of Saccharomyces cerevisiae could be measured in a range from 1 to 20 g L-1. In addition, the sensor can compensate for a change in the conductivity of the suspension of 5 to 15 mS cm-1. Moreover, the two-photon cross-linking of hydroxyethyl starch methacrylate hydrogel, which has been studied in detail, recommends itself for even much broader sensing applications in miniaturized bioreactors and biosensors.

High-Resolution Structuring of Silica-Based Nanocomposites for the Fabrication of Transparent Multicomponent Glasses with Adjustable Properties.

Silicate-based multicomponent glasses are of high interest for technical applications due to their tailored properties, such as an adaptable refractive index or coefficient of thermal expansion. However, the production of complex structured parts is associated with high effort, since glass components are usually shaped from high-temperature melts with subsequent mechanical or chemical postprocessing. Here for the first time the fabrication of binary and ternary multicomponent glasses using doped nanocomposites based on silica nanoparticles and photocurable metal oxide precursors as part of the binder matrix is presented. The doped nanocomposites are structured in high resolution using UV-casting and additive manufacturing techniques, such as stereolithography and two-photon lithography. Subsequently, the composites are thermally converted into transparent glass. By incorporating titanium oxide, germanium oxide, or zirconium dioxide into the silicate glass network, multicomponent glasses are fabricated with an adjustable refractive index nD between 1.4584-1.4832 and an Abbe number V of 53.85-61.13. It is further demonstrated that by incorporating 7 wt% titanium oxide, glasses with ultralow thermal expansion can be fabricated with so far unseen complexity. These novel materials enable for the first time high-precision lithographic structuring of multicomponent silica glasses with applications from optics and photonics, semiconductors as well as sensors.

Bone-Mimetic Osteon Microtopographies on Poly-ε-Caprolactone Enhance the Osteogenic Potential of Human Mesenchymal Stem Cells.

The attributes of implant surfaces are pivotal for successful osseointegration. Among surface engineering strategies, microtopography stands out as a promising approach to promote early cellular interactions. This study aims to design and craft a novel biomimetic osteon-like surface modification and to compare its impact on human mesenchymal stem cells (hMSCs) with four established topographies: blank, inverted pyramids, protrusions, and grooves. Poly-ε-caprolactone samples are fabricated using 2-photon-polymerization and soft lithography, prior to analysis via scanning electron microscopy (SEM), water contact angle (WCA), and protein adsorption assays. Additionally, cellular responses including cell attachment, proliferation, morphology, cytoskeletal organization, and osteogenic differentiation potential are evaluated. SEM confirms the successful fabrication of microtopographies, with minimal effect on WCA and protein adsorption. Cell attachment experiments demonstrate a significant increase on the osteon-like structure, being three times higher than on the blank. Proliferation assays indicate a fourfold increase with osteon-like microtopography compared to the blank, while ALP activity is notably elevated with osteon-like microtopography at days 7 (threefold increase over blank) and 14 (fivefold increase over blank). In conclusion, the novel biomimetic osteon-like structure demonstrates favorable responses from hMSCs, suggesting potential for promoting successful implant integration in vivo.

3D-printed ultra-small Brownian viscometers.

Measuring viscosity in volumes smaller than a microliter is a challenging endeavor. A new type of microscopic viscometers is presented to assess the viscosity of Newtonian liquids. Micron-sized flexible polymer cantilevers are created by two-photon polymerization direct laser writing. Because of the low stiffness and high elasticity of the polymer material the microcantilevers exhibit pronounced Brownian motion when submerged in a liquid medium. By imaging the cantilever's spherically shaped end, these fluctuations can be tracked with high accuracy. The hydrodynamic resistance of the microviscometer is determined by fitting the power spectral density of the measured fluctuations with a theoretical frequency dependence. Validation measurements in water-glycerol mixtures with known viscosities reveal excellent linearity of the hydrodynamic resistance to viscosity, allowing for a simple linear calibration. The stand-alone viscometer structures have a characteristic size of a few tens of microns and only require a very basic external instrumentation in the form of microscopic imaging at moderate framerates (~ 100 fps). Thus, our results point to a practical and simple to use ultra-low volume viscometer that can be integrated into lab-on-a-chip devices.

Femtosecond laser 3D printed micro objective lens for ultrathin fiber endoscope.

The most important optical component in an optical fiber endoscope is its objective lens. To achieve a high imaging performance level, the development of an ultra-compact objective lens is thus the key to an ultra-thin optical fiber endoscope. In this work, we use femtosecond laser 3D printing to develop a series of micro objective lenses with different optical designs. The imaging resolution and field-of-view performances of these printed micro objective lenses are investigated via both simulations and experiments. For the first time, multiple micro objective lenses with different fields of view are printed on the end face of a single imaging optical fiber, thus realizing the perfect integration of an optical fiber and objective lenses. This work demonstrates the considerable potential of femtosecond laser 3D printing in the fabrication of micro-optical systems and provides a reliable solution for the development of an ultrathin fiber endoscope.

Printing Green: Microalgae-Based Materials for 3D Printing with Light.

Microalgae have emerged as sustainable feedstocks due to their ability to fix CO2 during cultivation, rapid growth rates, and capability to produce a wide variety of metabolites. Several microalgae accumulate lipids in high concentrations, especially triglycerides, along with lipid-soluble, photoactive pigments such as chlorophylls and derivatives. Microalgae-derived triglycerides contain longer fatty acid chains with more double bonds on average than vegetable oils, allowing a higher degree of post-functionalization. Consequently, they are especially suitable as precursors for materials that can be used in 3D printing with light. This work presents the use of microalgae as "biofactories" to generate materials that can be further 3D printed in high resolution. Two taxonomically different strains -Odontella aurita (O. aurita, BEA0921B) and Tetraselmis striata (T. striata, BEA1102B)- are identified as suitable microalgae for this purpose. The extracts obtained from the microalgae (mainly triglycerides with chlorophyll derivatives) are functionalized with photopolymerizable groups and used directly as printable materials (inks) without the need for additional photoinitiators. The fabrication of complex 3D microstructures with sub-micron resolution is demonstrated. Notably, the 3D printed materials show biocompatibility. These findings open new possibilities for the next generation of sustainable, biobased, and biocompatible materials with great potential in life science applications.

3D Nanoprinting of Heterogeneous Metal Oxides with High Shape Fidelity.

3D nanoprinting can significantly enhance the performance of sensors, batteries, optoelectronic/microelectronic devices, etc. However, current 3D nanoprinting methods for metal oxides are suffering from three key issues including limited material applicability, serious shape distortion, and the difficulty of heterogeneous integration. This paper discovers a mechanism in which imidazole and acrylic acid synergistically coordinate with metal ions in water. Using the mechanism, this work develops a series of metal ion synergistic coordination water-soluble (MISCWS) resins for 3D nanoprinting of various metal oxides, including MnO2, Cr2O3, Co3O4, and ZnO, as well as heterogeneous structures of MnO2/NiO, Cr2O3/Al2O3, and ZnO/MgO. Besides, the synergistic coordination effect results in a 2.54-fold increase in inorganic mass fraction within the polymer, compared with previous works, which effectively mitigates the shape distortion of metal oxide microstructures. Based on this method, this work also demonstrates a 3D ZnO microsensor with a high sensitivity (1.113 million at 200 ppm NO2), surpassing the conventional 2D ZnO sensors by tenfold. The method yields high-fidelity 3D structures of heterogeneous metal oxides with nanoscale resolution, paving the way for applications such as sensing, micro-optics, energy storage, and microsystems.

Field Emission from Carbon Nanotubes on Titanium Nitride-Coated Planar and 3D-Printed Substrates.

Carbon nanotubes (CNTs) are well known for their outstanding field emission (FE) performance, facilitated by their unique combination of electrical, mechanical, and thermal properties. However, if the substrate of choice is a poor conductor, the electron supply towards the CNTs can be limited, restricting the FE current. Furthermore, ineffective heat dissipation can lead to emitter-substrate bond degradation, shortening the field emitters' lifetime. Herein, temperature-stable titanium nitride (TiN) was deposited by plasma-enhanced atomic layer deposition (PEALD) on different substrate types prior to the CNT growth. A turn-on field reduction of up to 59% was found for the emitters that were generated on TiN-coated bulk substrates instead of on pristine ones. This observation was attributed exclusively to the TiN layer as no significant change in the emitter morphology could be identified. The fabrication route and, consequently, improved FE properties were transferred from bulk substrates to free-standing, electrically insulating nanomembranes. Moreover, 3D-printed, polymeric microstructures were overcoated by atomic layer deposition (ALD) employing its high conformality. The results of our approach by combining ALD with CNT growth could assist the future fabrication of highly efficient field emitters on 3D scaffold structures regardless of the substrate material.

3D-Printed Ultracompact Multicore Fiber-Tip Probes for Simultaneous Measurement of Nanoforce and Temperature.

Optical fiber force sensing has attracted considerable interest in biological, materials science, micromanipulation, and medical applications owing to its compact and cost-efficient configuration. However, the glass fiber has an intrinsic high Young's modulus, resulting in force sensors being generally less sensitive. While hyperelastic polymer materials can be utilized to enhance the force sensitivity, the thermodynamic properties of the polymer may weaken the sensing accuracy and reliability. Herein, we demonstrate ultracompact three-dimensional (3D)-printed multicore fiber (MCF) tip probes for simultaneous measurement of nanoforce and temperature with high sensitivity. The sensor is highly sensitive to force-induced deformation due to the special geometric features of the polymer microcantilever, and the high-temperature sensitivity can be implemented through the poly(dimethylsiloxane) (PDMS) microcavity on the same fiber facet. Moreover, the sensitivities of the fiber interferometers are remarkably enhanced by introducing the optical analogue of the Vernier effect. Such a device exhibits a force sensitivity of 56.35 nm/µN, which is more than 103 times that of all-silica fiber force sensors. The PDMS microcavity provides a temperature sensitivity of 1.447 nm/°C, measuring the local temperature of the probe and compensating for temperature crosstalk of the force detection. The proposed compact MCF-tip sensor can simultaneously measure nanoforce and temperature with high sensitivity, facilitating multiparameter sensing in a restricted space environment and showing the potential in miniaturized all-fiber multiparameter sensors.

3D Directional Assembly of Liquid Crystal Molecules.

The precise construction of hierarchically long-range ordered structures using molecules as fundamental building blocks can fully harness their anisotropy and potential. However, the 3D, high-precision, and single-step directional assembly of molecules is a long-pending challenge. Here, a 3D directional molecular assembly strategy via femtosecond laser direct writing (FsLDW) is proposed and the feasibility of this approach using liquid crystal (LC) molecules as an illustrative example is demonstrated. The physical mechanism for femtosecond (fs) laser-induced assembly of LC molecules is investigated, and precise 3D arbitrary assembly of LC molecules is achieved by defining the discretized laser scanning pathway. Additionally, an LC-based Fresnel zone plate array with polarization selection and colorization imaging functions is fabricated to further illustrate the potential of this method. This study not only introduces a 3D high-resolution alignment method for LC-based functional devices but also establishes a universal protocol for the precise 3D directional assembly of anisotropic molecules.

Structural Colored Based Humidity Sensor Consisting of High Resolution 3D Printed Photonic Crystal Coated with Ultrathin Responsive Hydrogels.

Today, humidity sensors have become an integral part of the daily lives. In particular, humidity sensors using an electronic measuring principle have become the standard. Although these sensors have proven to be a stable measurement method, they have some disadvantages, such as their long response time or the danger of using them in explosive environments. This work introduces photonic crystals as an alternative optical measurement approach. The novel technology of ultra-fast two-photon polymerisation printing is combined with a thin-film deposition process, namely iCVD. This allows to print large area high-precision 3D templates, which are subsequently coated with a humidity responsive hydrogel thin film (p(HEMA) of 20 nm.The limits of 2PP technology are being pushed allowing the production ofs table and periodic large-area 3D structures. The flexible customization of hydrogels for ambient conditions make them exceptionally promising for a wide range of sensing applications. Additionally, optical methods for measuring humidity seem to be an excellent alternative to overcome the limitations for current state of the art humidity sensors. The optical detection of changes in ambient air humidity is achieved by observing color changes of the printed structure within the visible wavelength range.

Dimensionality Matters: Exploiting UV-Photopatterned 2D and Two-Photon-Printed 2.5D Contact Guidance Cues to Control Corneal Fibroblast Behavior and Collagen Deposition.

In the event of disease or injury, restoration of the native organization of cells and extracellular matrix is crucial for regaining tissue functionality. In the cornea, a highly organized collagenous tissue, keratocytes can align along the anisotropy of the physical microenvironment, providing a blueprint for guiding the organization of the collagenous matrix. Inspired by this physiological process, anisotropic contact guidance cues have been employed to steer the alignment of keratocytes as a first step to engineer in vitro cornea-like tissues. Despite promising results, two major hurdles must still be overcome to advance the field. First, there is an enormous design space to be explored in optimizing cellular contact guidance in three dimensions. Second, the role of contact guidance cues in directing the long-term deposition and organization of extracellular matrix proteins remains unknown. To address these challenges, here we combined two microengineering strategies-UV-based protein patterning (2D) and two-photon polymerization of topographies (2.5D)-to create a library of anisotropic contact guidance cues with systematically varying height (H, 0 µm ≤ H ≤ 20 µm) and width (W, 5 µm ≤ W ≤ 100 µm). With this unique approach, we found that, in the short term (24 h), the orientation and morphology of primary human fibroblastic keratocytes were critically determined not only by the pattern width, but also by the height of the contact guidance cues. Upon extended 7-day cultures, keratocytes were shown to produce a dense, fibrous collagen network along the direction of the contact guidance cues. Moreover, increasing the heights also increased the aligned fraction of deposited collagen and the contact guidance response of cells, all whilst the cells maintained the fibroblastic keratocyte phenotype. Our study thus reveals the importance of dimensionality of the physical microenvironment in steering both cellular organization and the formation of aligned, collagenous tissues.

4D Printed Soft Microactuator for Particle Manipulation via Surrounding Medium Variation.

Soft actuators have assumed vital roles in a diverse number of research and application fields, driving innovation and transformative advancements. Using 3D molding of smart materials and combining these materials through structural design strategies, a single soft actuator can achieve multiple functions. However, it is still challenging to realize soft actuators that possess high environmental adaptability while capable of different tasks. Here, the response threshold of a soft actuator is modulated by precisely tuning the ratio of stimulus-responsive groups in hydrogels. By combining a heterogeneous bilayer membrane structure and in situ multimaterial printing, the obtained soft actuator deformed in response to changes in the surrounding medium. The response medium is suitable for both biotic and abiotic environments, and the response rate is fast. By changing the surrounding medium, the precise capture, manipulation, and release of micron-sized particles of different diameters in 3D are realized. In addition, static capture of a single red blood cell is realized using biologically responsive medium changes. Finally, the experimental results are well predicted using finite element analysis. It is believed that with further optimization of the structure size and autonomous navigation platform, the proposed soft microactuator has significant potential to function as an easy-to-manipulate multifunctional robot.

A Multi-foci Sparse-Aperture Metalens.

Multi-foci lenses are essential components for optical communications, virtual reality display and microscopy, yet the bulkiness of conventional counterparts has significantly hindered their widespread applications. Benefiting from the unprecedented capability of metasurfaces in light modulation, metalenses are able to provide multi-foci functionality with a more compact footprint. However, achieving imaging quality comparable to that of corresponding single-foci metalenses at each focal point poses a challenge for existing multi-foci metalenses. Here, a polarization-independent all-dielectric multi-foci metalens is proposed and experimentally demonstrated by spatially integrating single-foci optical sparse-aperture sub-metalenses. Such design enables the metalens to generate multiple focal points, while maintaining the ability to capture target information comparable to that of a single-foci metalens. The proposed multi-foci metalens is composed of square-nanohole units array fabricated by two-photon polymerization. The focusing characteristic and imaging capability are demonstrated upon the illumination of an unpolarized light beam. This work finds a novel route toward multi-foci metalenses and may open a new avenue for dealing with the trade-off between multi-foci functionality and high-quality imaging performance.

3D micro/nano hydrogel structures fabricated by two-photon polymerization for biomedical applications.

Hydrogels are three-dimensional natural or synthetic cross-linked networks composed of polymer chains formed by hydrophilic monomers. Due to the ability to simulate many properties of natural extracellular matrix, hydrogels have been widely used in the biomedical field. Hydrogels can be obtained through a variety of polymerization strategies such as heating and redox. However, photochemistry is one of the most interesting methods for researchers in this field. Gelatin-methacryloyl (GelMA) inherits the biological activity of gelatin and has become one of the gold standards in the field of biomaterials. GelMA, as a photopolymerizable hydrogel precursor, can be used to fabricate 3D porous structures for biomedical applications through two-photon polymerization. We report a new formulation of GelMA-based photoresist and used it to manufacture a series of two-photon polymerization structures, with a maximum resolution less than 120 nm. The influence of process parameters on 3D structures manufacturing is studied by adjusting the scanning speed, laser power, and layer spacing values in two-photon polymerization processing. In vitro biological tests show that the 3D hydrogel produced by two-photon polymerization in this paper is biocompatible and suitable for MC3T3-E1 cell.

FABRICATION OF MULTILUMEN MICROFLUIDIC TUBING FOR EX SITU DIRECT LASER WRITING.

Among the numerous additive manufacturing or "three-dimensional (3D) printing" techniques, two-photon Direct Laser Writing (DLW) is distinctively suited for applications that demand high geometric versatility with micron-to-submicron-scale feature resolutions. Recently, "ex situ DLW (esDLW)" has emerged as a powerful approach for printing 3D microfluidic structures directly atop meso/macroscale fluidic tubing that can be manipulated by hand; however, difficulties in creating custom esDLW-compatible multilumen tubing at such scales has hindered progress. To address this impediment, here we introduce a novel methodology for fabricating submillimeter multilumen tubing for esDLW 3D printing. Preliminary fabrication results demonstrate the utility of the presented strategy for resolving 743 µm-in-diameter tubing with three lumens-each with an inner diameter (ID) of 80 µm. Experimental results not only revealed independent flow of discrete fluorescently labelled fluids through each of the three lumens, but also effective esDLW-printing of a demonstrative 3D "MEMS" microstructure atop the tubing. These results suggest that the presented approach could offer a promising pathway to enable geometrically sophisticated microfluidic systems to be 3D printed with input and/or output ports fully sealed to multiple, distinct lumens of fluidic tubing for emerging applications in fields ranging from drug delivery and medical diagnostics to soft surgical robotics.

TOWARD CONTROLLED-RELEASE DRUG DELIVERY MICROCARRIERS ENABLED BY DIRECT LASER WRITING 3D PRINTING.

Controlled-release, and especially long-acting, drug delivery systems hold promise for improving treatments for numerous medical conditions. Previously, we reported an additive manufacturing or "three-dimensional (3D) printing" approach for fabricating liquid-core-shell-cap microcarriers comprising standard photoresists. Here we explore the potential to extend this strategy to achieve microcarriers comprising biodegradable materials as a new pathway to controlled-release drug delivery options. Specifically, we investigate the use of "Two-Photon Direct Laser Writing (DLW)" as a means to 3D print microcarriers composed of: (i) a bottle-shaped "shell" with an orifice, (ii) an aqueous liquid "core", and (iii) a biodegradable "cap". The cap, which is DLW-printed directly onto the shell's orifice, is designed to degrade over time in the body-e.g., with degradation time proportional to cap thickness-to ultimately facilitate release of the liquid core at desired time points. Fabrication results based on the use of a biodegradable poly(ethylene glycol) diacrylate (PEGDA) photomaterial for the cap revealed that shell designs incorporating microfluidic obstruction structures appeared to limit undesired entry of the liquid-phase PEGDA into the shell (i.e., directly preceding cap printing), thereby resulting in improved retention of the liquid core after completion of the cap printing process. These results mark an important first step toward evaluating the utility of the presented DLW 3D printing strategy for possible drug delivery applications.

A 3D-MICROPRINTED COAXIAL NOZZLE FOR FABRICATING LONG, FLEXIBLE MICROFLUIDIC TUBING.

A variety of emerging applications, particularly those in medical and soft robotics fields, are predicated on the ability to fabricate long, flexible meso/microfluidic tubing with high customization. To address this need, here we present a hybrid additive manufacturing (or "three-dimensional (3D) printing") strategy that involves three key steps: (i) using the "Vat Photopolymerization (VPP) technique, "Liquid-Crystal Display (LCD)" 3D printing to print a bulk microfluidic device with three inlets and three concentric outlets; (ii) using "Two-Photon Direct Laser Writing (DLW)" to 3D microprint a coaxial nozzle directly atop the concentric outlets of the bulk microdevice, and then (iii) extruding paraffin oil and a liquid-phase photocurable resin through the coaxial nozzle and into a polydimethylsiloxane (PDMS) channel for UV exposure, ultimately producing the desired tubing. In addition to fabricating the resulting tubing-composed of polymerized photomaterial-at arbitrary lengths (e.g., > 10 cm), the distinct input pressures can be adjusted to tune the inner diameter (ID) and outer diameter (OD) of the fabricated tubing. For example, experimental results revealed that increasing the driving pressure of the liquid-phase photomaterial from 50 kPa to 100 kPa led to fluidic tubing with IDs and ODs of 291±99 µm and 546±76 µm up to 741±31 µm and 888±39 µm, respectively. Furthermore, preliminary results for DLW-printing a microfluidic "M" structure directly atop the tubing suggest that the tubing could be used for "ex situ DLW (esDLW)" fabrication, which would further enhance the utility of the tubing.