Symposium SM08—Next-Generation Materials and Technologies for 3D Printing and Bioprinting

Design for Additive Manufacturing (AM) is widely acknowledged as a severe barrier to the progression of the technology. In terms of biomaterials, whilst AM provides unique opportunities for the design of geometries and material compositions, there remains a persistent problem in the trade-off between the desired mechanical properties and the material properties [1]. To overcome this challenge, a radically new interdisciplinary approach to the development of AM systems and design techniques for biomaterials is required. Rather than a bioinspired method towards designing a single product, this research outlines how the exploration of developmental biology can expand the frontiers of AM systems and design techniques [2].

The majority of AM techniques increase the part size through incremental fusion of the material. Considering the development of the human in-utero; life begins as a single cell, which eventually forms tissues and organs, growing and functioning in synergy. The essence of foetal development is a very sophisticated AM process; in-utero growth demonstrates intrinsically intertwined materials, design and manufacturing [2]. However, this synergy between materials, design and manufacturing is not reflected in AM.

The development of the foetus is fundamentally dependent on varying spatial and temporal stimuli through the pregnancy. Analogical reasoning between foetal development and AM highlights a complete lack of any tailored process parameters during manufacturing. Process parameters are currently predefined and remain constant. Yet the process parameters underpin the properties of the material. Temporal DfAM, the design of process parameters during an AM build, is proposed as a method of designing the properties of a material through a single part [3]. Whilst this approach has been demonstrated computationally, its application to biomaterials is novel.

Poly(vinyl alcohol) (PVA) cryogel is a biocompatible, synthetic hydrogel that can be physically cross-linked at sub-zero temperatures. The mechanical properties of PVA are tailored through concentration, molecular weight and the number and temperature profile of freezing and thawing cycles. This makes it a widely applicable biomaterial as a component of tissue constructs and diagnostic phantoms. A novel cryogenic 3D printer has been developed to facilitate the AM of PVA. In doing so, the geometric complexity of the meso and micro-structure of the material is vastly increased.

This presentation will demonstrate the application of the novel design technique TDfAM to a cryogenic 3D printing platform for PVA. Changing the fixed process parameters of cryogenic 3D printing leads to a change in the mechanical properties of the PVA. Thus, TDfAM, the variation of process parameters during an AM build, will be explored with respect to the local and global material heterogeneity and mechanical properties. This experimental work will be validated through computational simulations.

The limitations of the TDfAM technique and cryogenic 3D printing of PVA will be discussed with respect to the current barriers to their industrial and clinical translation. Finally, we will outline the next steps of this unique research trajectory, exploring new dimensions of developmental biology and how we can develop current research and extract more novel innovations in AM design and manufacturing systems.

[1] S. Miramini et al. (2020) Journal of the Mechanical Behaviour of Biomedical Materials, 103: 103544
[2] L.E.J. Thomas-Seale et al. (2019) Cogent Engineering, 6:1662631
[3] S. Saliba et al. (2020) International Journal of Advanced Manufacturing Technology, 106: 3849-3857

Two photon polymerization is a 3D printing process that relies on the use of ultrashort laser pulses to selectively polymerize and solidify photosensitive materials. The quadratic character of the two photon absorption probability and the well-defined polymerization threshold of this process allow for the diffraction limit to be exceeded and structures with features below one micrometer to be prepared. Two photon polymerization has recently been to fabricate many types of microstructured and nanostructured medical devices out of biocompatible inorganic-organic hybrid materials (such as zirconium oxide hybrid materials) and polymers for medical applications. The use of biocompatible photoinitiators, including a combination of riboflavin and triethanolamine, for two photon polymerization will be considered. Integration of electrochemical sensors with two photon polymerization-processed microneedles will be discussed. Assessment of the biocompatibility of two photon polymerization-processed materials using in vitro biological studies will be discussed. In addition, application-specific studies of two photon polymerization-processed medical devices for biosensing and drug delivery, including in vivo studies, will be described. Our results indicate that two photon polymerization provides many benefits for fabricating medical devices with small-scale features and unique drug delivery and biosensing functionalities.

Inkjet printing is a powerful enabling technology for formation of structures at micro and nanoscale for current mosto advanced fileds of appplications. As it is a direct fabrication approach, it has the undisputed advantage to avoid any masks, molds or lithography steps. Furthermore, in addition to this great flexibility, recently it has been demonstrated the capability of inkjet printing approaches to reach very high spatial resolution, down to nanoscale and over large area. Direct printing of micro and nano structures has been proofed with a variety of different inks and different types of materials. Different technologies exist, such as thermal jetting, piezo-based jetting, Electrohydrodynamic (EHD) jetting, syringe-based pumping or simply contact-dispensing printing. Each of the above-mentioned technologies is based on liquid media to be delivered in the jet printing process. Moreover, most of the existing methods include needles for liquid dispensing. Unfortunately, the use of needles has some severe limitations such as clogging and cross contamination. Here it will be presented some recent developments about a novel and special platform based on EHD jetting driven by a pyroelectric effect activated in a bulk ferroelectric crystal. Waht we named Pyro-EHD exhibits significant potentialities in allowing liquid dispensing and printing for a number of applications ranging from microelectronics to biotechnologies. One of the interesting features is that pyro-EHD can easily work in nozzle-less modality thus avoiding needles and consequently the related clogging drawbacks. The pyro-EHD printer approach usually dispenses liquids from a liquid reservoir that can be a sessile droplet or even the free surface of a liquid. The high potentials easily attainable through the ignition of the pyroelectric effect allow to manouvering and dispense and print also high viscous polymers and liquids dropltes. Description of the pyro-EHD will be provided. The results obtained will be shown and discussed. Several examples will be illustrated for direct printing of liquids under both single and multiphase forms that can find useful exploitation in a multiplicity of fields from microelectronics to biomedical applications.
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[3] Ferraro, P., Grilli, S., Miccio, L., & Vespini, V. (2008). Wettability patterning of lithium niobate substrate by modulating pyroelectric effect to form microarray of sessile droplets. Applied Physics Letters, 92(21), 213107.
[4] Coppola, S., Vespini, V., Nasti, G., Gennari, O., Grilli, S., Ventre, M., ... & Ferraro, P. (2014). Tethered pyro-electrohydrodynamic spinning for patterning well-ordered structures at micro-and nanoscale. Chemistry of Materials, 26(11), 3357-3360.
[5] Nasti, G., Coppola, S., Vespini, V., Grilli, S., Vettoliere, A., Granata, C., & Ferraro, P. (2020). Pyroelectric tweezers for handling liquid unit volumes. Advanced Intelligent Systems, 2000044.
[6] Coppola, S., Nasti, G., Vespini, V., & Ferraro, P. (2020). Layered 3D printing by tethered pyro-electrospinning. Advances in Polymer Technology, 2020.
[7] Vecchione, R., Coppola, S., Esposito, E., Casale, C., Vespini, V., Grilli, S., ... & Netti, P. A. (2014). Electro drawn drug loaded biodegradable polymer microneedles as a viable route to hypodermic injection. Advanced Functional Materials, 24(23), 3515-3523.

In the last decades, advances in bone tissue engineering mainly based on osteoinduction and on stem cell research. Only recently, new efforts by others and us focused on the micro- and nanoarchitecture needed to improve and accelerate bone regeneration. By the use of additive manufacturing, libraries of diverse microarchitectures were produced and tested to identify the ideal pore size or rod distance for osteoconduction to occur. Presently, we try to elucidate the dependency of osteoconduction on microporosity and expand our view on micro- and nanoarchitecture of bone substitutes for optimal bone augmentation.
For the production of scaffolds, we applied for titanium-based scaffolds selective laser melting and for ceramics the CeraFab 7500 from Lithoz, a lithography-based additive manufacturing machine. As in vivo test model, we used a calvarial defect and a bone augmentation model in rabbits. The histomorphometric analysis showed that bone formation was significantly increased with pores between 0.7-1.2 mm in diameter. Moreover, microporosity appeared to be a strong driver of osteoconduction and influenced osteoclastic degradation. Best microarchitecture for osteoconduction and bone augmentation are different.
In essence, additive manufacturing enabled us to generate libraries of microarchitectures to search for the most osteoconductive microarchitecture and the ideal microarchitecture for bone augmentation purposes. Moreover, additive manufacturing appears as a promising tool for the production of personalized bone tissue engineering scaffolds to be used in cranio-maxillofacial surgery, dentistry, and orthopedics.

Additive manufacturing of hydrogels or elastomers can be used to fabricate microfluidic channels with biomimetic functionality. For instance, atomically thin sheets of nanomaterials can augment organic polymers with enhanced mechanical and physical functionalities. These 3D printed structures can further exhibit stimuli-responsive behaviors such as actuation, self-adhesion, or degradation. Here, we present recent results on light-directed 3D printing of next generation materials using covalent and ionic crosslinking. First, we explore the use of versatile alginate hydrogels with graphene oxide to pattern collective cell migration, tune stimuli-responsive behaviors, and repel oil in seawater like conditions. Second, we demonstrate double network hydrogels that are stimuli-responsive and self-adhesive as a simple “do-it-yourself” construction set for soft machines and microfluidic devices. This approach enables “plug-and-play” hydrogel parts for ionic soft machines that emulate actuation, sensing, and fluid transport in living systems. Finally, we investigate the use of 3D printed elastomers for microfluidic devices to achieve perfusion of tissue fragments for preclinical testing of immunotherapies. Overall, 3D printing enables multiscale integration of nanomaterial and polymeric building blocks into bespoke larger scale structures for advanced materials and biomedicine.

The tumor microenvironment (TME) is a dynamic cellular and biochemical system, integrating multiple positive and negative biological feedback loops that influence cell physiology. Cell-extracellular matrix, secreted factor-cell signaling pathways, and cell-cell interactions are known factors in tumorigenicity and drivers of therapeutic resistance. There is, therefore, an urgent need to develop clinically relevant in vitro platforms comprising epithelial and stromal cells in an extracellular matrix mimicking microenvironment. Three-dimensional (3D) cell culture and co-culture of cancer cells have demonstrated that creating physiologically representative tumor microenvironment in vitro models enables patient-specific experiments to be performed. Typically, these are performed by casting cells within a droplet of Matrigel, or commercially available biocompatible hydrogels, promoting the self-assembly of multi-cellular organoids. While these systems provide a more relevant microenvironment than monolayers, the resulting organoids have limited user input over parameters such as the initial location of different cell types.

We have recently developed extracellular matrix mimicking hydrogel biomaterials capable of hosting cells, and subsequently, being extruded to form complex in vitro tumor models. Our lab has previously demonstrated that composite materials constituted of sodium alginate, a seaweed-derived polysaccharide, and gelatin, a bovine or porcine-derived denatured collagen, can be tuned to recapitulate the mechanical properties of soft tissues while keeping their biocompatibility and printability. Here, we report a bioink prepared by decellularizing tissue and incorporating sodium alginate and gelatin at controlled weight percentages as rheological modifiers to reinforce and positively impact the composite material's mechanical integrity. dECM contains structural proteins, glycosaminoglycans, and growth factors preserved from the tissue of origin. Gelatin provides mechanical stability during the printing process, and crosslinked-alginate chains maintain sample integrity in long-term cell culture conditions.

We have demonstrated this methodology to create breast and head and neck tumor models using immortalized cell lines and, more recently, lung, gastric, and breast models derived from patient tumor samples to create precision interventional schemes. Of particular interest is the ability to build complex 3D geometric structures using soft materials (bioinks) that recapitulate the mechanical properties such as the stiffness of solid tumors and their microenvironment. Bioprinting permits more complex geometric matrices to be printed with high cell density, control over cell location, and viability. Bioprinted models can be created directly with precise reproducibility from cell-hydrogel suspensions with high throughput and scalability directly on to well plate platforms. In collaboration with the Sangwan and Ferri labs at the Goodman Cancer Research Centre at McGill University, we have further explored using biobanked tissue samples to develop personalized tumor models.

Three-dimensional hierarchical structures with controllable mechanical properties are expected to have wide range of applications. In general, material properties depend on the building block and it remains challenging to develop materials with dramatically different mechanical properties from the same type of material. In this presentation, cellulose was demonstrated for 3D printing of two types of structure. In one case, a lightweight (∼90 mg/cm³) and super-strong (16.6 MPa compressive Young’s modulus) honeycomb structure is constructed by 3D printing of cellulose ink dissolved in NaOH/urea solution. The 3D printed cellulose structure demonstrated switchable high elasticity (to withstand varied repetitive elastic deformation) at the wet state and high rigidity (to support over 15,800 of its own weight) at dry state. In another case, we demonstrated a compressible and superelastic 3D printed structure based on cellulose nanofibrils. The 3D structure showed superb elasticity (over 91% strain recovery after 500 cycles of compressive test) and compressibility (up to 90% compressive strain). In addition, with the incorporation of salt, the 3D printed CNF structure was demonstrated to serve as pressure sensor with high pressure sensitivity of 0.337 kPa^-1 at 43% relative humidity. These results indicate that the mechanical properties of the 3D printed cellulose structure can be facilely controlled by the inter-fibril interactions, leading to either super-strong or super-elastic materials properties.

Bio-sourced materials that can replace existing petroleum-based materials are an integral component of sustainability. Moreover, biomaterials with greater complexity and functionality will be required to meet the demands for the full spectrum of applications from aerospace to medicine. The unique features of proteins (such as the mechanical properties of spider silk) have inspired the development of protein-based plastics, but their application is limited by poor processability and limitations in mechanical performance.
Both 3D and 4D additive manufacturing (AM) processes that utilize vat photopolymerization have tremendous potential in industrial manufacturing for the future production of parts and supplies. Naturally occurring polymers often require modification with photocurable functionalities to become printable. Most notably, methacryl groups have been introduced onto the amines of biopolymers to afford polymerizable materials like gelatin methacrylamide (Gel-MA), silk fibroin methacrylate (Sil-MA), and methacrylated bovine serum albumin (MA-BSA). While 4D printed structures that undergo chemical or physical changes in response to their environment are often inspired by nature, there are relatively few examples that use bio-sourced and biodegradable materials.
Both structural and globular proteins are known as macromolecules that can introduce material plasticity or elasticity based on the unfolding or disassembly of the proteins. Proteins can be unravelled reversibly or irreversibly upon the application of a tensile force, thus, the secondary and tertiary structure of proteins have been utilized to achieve biomaterials with unique combinations of extensibility, strength, and resilience. More recently, BSA was incorporated into hydrogel networks to provide an energy dissipation mechanism via protein unfolding. However, there has not been a demonstration of a protein-based material that exhibits plasticity due to unfolding, with a corresponding shape recovery back to its original form.
We formulated a simple aqueous-based resin for laser-scanning SLA printing based on BSA in a photocurable resin. BSA is an abundant globular protein with excellent aqueous solubility and low viscosity at concentrations up to 30 wt%. Furthermore, we demonstrated that the native conformation of these globular proteins is largely retained in the 3D printed constructs, and that each protein molecule possesses a “stored length” that could be revealed during mechanical deformation (extension or compression) of the 3D bioplastic objects. Given the large molecular weight of BSA (66 kDa), the potential for unfolding individual protein chains can be significant, as exhibited by the plasticity observed in these thermoset materials. We expect this strategy – wherein globular proteins are utilized to afford 3D printed bioplastics with mechanical properties that utilize the stored length of these macromolecules – is broadly applicable to other forms of vat photopolymerization to create constructs that can create a closed loop life cycle with bioplastics. In short, we introduced a bio-sourced resin for additive manufacturing that can be used to fabricate in a rapid manner complex elements for a wide range of applications, with a very interesting stimuli response of the printed parts, due to the conformational changes of proteins at the molecular level.

Luminescent lanthanide doped nanoparticles have received significant attention due to their fascinating optical properties. At the core of this interest is their ability to convert near-infrared (NIR) excitation light (typically 980 or 800 nm) to higher energies spanning the ultraviolet (UV), visible and higher energy NIR regions of the spectrum through a multiphoton process known as upconversion. Upconversion differs from conventional multiphoton excitation in other materials where no real intermediate states are present necessitating the use of ultrafast lasers (in the femtosecond regime) for simultaneous excitation to the upper emitting state. The lanthanide ions possess a multitude of 4f electronic energy states that have long lifetimes (micro- to millisecond) thus act as population reservoirs in the upconversion process. Hence, upconversion occurs through real, long-lived intermediate states through a sequential photon absorption process. This eliminates the need for ultrafast excitation and as a result, upconversion can be observed using inexpensive, continuous wave diode lasers. Upconversion luminescence can be exploited for a number of applications in nanomedicine, theranostics, photovoltaics, photocatalysis, as well as many others.

In this presentation, we will discuss the synthesis of these nanoparticles, establish how changing their nanoscale architecture can affect their luminescence properties and upconversion, as well as discuss their applications in nanomedicine, theranostics and potentially in 3D printing.

Fused Deposition Modeling (FDM) has been adapted to metals using filaments made of metal particle reinforced polymer matrix composites (PMC). However, there is still a fundamental gap in understanding how its melt suspension rheology affects the ability to extrude and print filaments continuously without jamming. In this correlative study, a framework for studying rheological scaling is presented as function of increasing metal content (i.e. gas atomized NiCu) in a thermoplastic binder (i.e. PLA), and it was shown that, even with high zero-shear viscosities (more than 1x10⁵ Pa-s), high metal content composites showed a dramatic shear thinning behavior and hence were extrudable at a content of 54 vol% of metal. Extruded filaments were re-melted and submitted to flow sweep rheology test. The zero-shear viscosity scaling with metal content fits well with the existing theoretical model of Krieger and Daughtery, but it was not sufficient to predict the extrudability of the filament due to its complex shear thinning behavior. Additionally, the filaments displayed premature slippage onset at <10 s^-1 which is typically below those observed inside of the die during extrusion indicating potential change in the flow regime during extrusion. The homogeneity of composite filaments was also analyzed in global scale (by doing thermogravimetric analysis of the 12 ft continuously extruded filaments along the length to find out the metal vol%) and local scale (by analyzing a small cross-section in X-ray micro-tomography). It was verified that the filament content was uniform up in filaments with a maximum metal content of 54%. This strategy of homogeneously packing gas atomized metal spheres at higher metal content, gave us the insights on how to scale and modulate the melt rheology to create consistently extrudable and uniform dense metal filaments easily without using any kind of additives.

Nonthermal plasmas have proven to be an effective method for the synthesis of high-quality nanoparticles. The nonthermal plasma environment allows for crystallization despite the low temperature, suppression of particle agglomeration, and control over particle size through manipulation of the synthesis conditions. Typical nonthermal plasma reactors operating under vacuum are fairly large and costly. By operating at atmospheric pressure and miniaturizing the reactor, costs of the system are reduced and the ability to scale the process is enhanced. Reactors often benefit from confinement to the microscale at atmospheric pressure, this reduction in size lends itself well to act as a deposition head for additive manufacturing processes.
Here we present our work on the synthesis and deposition of nanoparticles into ordered patterns and structures. A 13.56 MHz RF power supply was used to generate a plasma in a glass capillary tube between two external ring electrodes at atmospheric pressure. Supplied to the reactor was a precursor gas of silane and a background gas of argon for the synthesis of silicon nanoparticles. The reactor was attached to a computer-controlled gantry and stage for the controlled deposition of the nanoparticles. The gas phase synthesis method inherently produces an aerosol of nanoparticles without the need of an atomizer as is required in typical aerosol jet printing processes (AJP). Typical AJP requires the use of an ink for atomization whose properties must be precisely to known to reduce losses during transportation and deposition. From TEM and XRD analysis we have been able to deposit crystalline silicon nanoparticles with tunable particle sizes approaching the Bohr-exciton radius of silicon that also exhibit a fairly narrow size distribution without the use of an ink. Further by varying synthesis conditions during deposition we have been able to print materials whose properties vary spatially along the deposition. Further the particles were deposited with linewidths as small as 100 microns and layer thicknesses of 1.5 microns.

Hydrogels are recognized as one of the most promising functional materials, as it is dynamic, tunable, biocompatible, biodegradable and able to encapsulate large water content. Owing to these unique characteristics, hydrogel based systems are revolutionizing many applications in solar energy water nexus, biomedical engineering, drug delivery and soft electronics. Hydrogel materials endow an intriguing activation phenomenon on water molecules confined in hydrogel meshes. By altering resin formulation and crosslinking density of photocrosslinked hydrogels, we are able to manipulate the water states within the hydrogel. The water states as well as the degree of photopolymerization are characterized by confocal Raman microscopy. Moreover, 3D microstructures of responsive hydrogels are fabricated using high resolution micro projection stereolithography. The transition from 3D hydrogel microstructures to 4D is studied during its response to external stimuli such as temperature and moisture, and its swelling dynamics is monitored through in situ condensation experiments using the environmental scanning electron microscopy. Finally, by fabricating functional hydrogel devices with tunable water states, we aim at developing high resolution 3D printing technologies for broad energy and sustainability applications.

Wear resistance and microstructure of potential prosthetic hip and joint bioimplant material system, a reactor blend of high density polyethylene (HDPE) and 10 wt% ultra high molecular weight polyethylene (UHMWPE), 3D printed using fused deposition modeling (FDM) with different printing speeds and printing orientations have been studied using microscratch tester and wide angle X-ray diffraction respectively and compared with those of injection molded samples. The microstructure has been represented in terms of texture components using pole figures and orientation distribution functions (ODFs). This study investigates how 3D printing parameters affect texture and how global combination of the texture components prevalent in different plastic regimes engender material responses to wear resistance. The effect of abrasion and change in wear resistance of the bioimplant material (HDPE+10wt%UHMWPE) system have been explored by varying the loads applied, scratch directions, scratch speeds and the number of recurring scratches within the microscratch tests. This study of correlation between wear resistance and texture suggests that improved wear resistance in the (HDPE+10wt%UHMWPE) system can be achieved by texturing the bioimplant material in the loading direction.
Keywords: High density polyethylene; Ultra high molecular weight polyethylene; 3D printing; Fused Deposition Modeling (FDM); Wear resistance; Texture; Bioimplant material;

Magnetoelectric materials are an important class of multifunctional materials with applications spanning from wearable electronics to space exploration. The current state-of-the-art in processing this exciting class of materials heavily rely on vacuum-based methods, increasing the complexity and hindering their translational potential. Magnetoelectric materials couple, intrinsically or extrinsically, the magnetic and electric energies through several mediators, including strain, charge, and spin, making them imperative for straintronics, optoelectrics, and spintronics applications. The objective of this research is to report a novel processing method of polymer-based magnetoelectric composite materials using additive manufacturing technique. The latter is parametrically different than the state-of-the-art, resulting in reducing cost, increasing yield, and streamlining processing of multifunctional materials. The approach is realized composites of electroactive polymer with large piezoelectric properties and magnetic particles using digital light processing technique (DLP), whereas the application of an electric field across the surface results in change in magnetization while a magnetic field changes the state of polarization, hence the magnetoelectric coupling. Notably, previous research has shown that the suspension of nanoscale magnetic particles increased the dielectric properties of the electroactive phase without affecting its piezoelectric response. A photopolymer resin is use as the binder, which is activated by the light source in the printer within the ultraviolet spectrum. Additive manufacturing with DLP technique produces lightweight and nearly isotropic structures with high geometrical accuracy, in comparison with other methods such as material extrusion. Samples with different volume fractions of magnetic particles and electroactive polymers were fabricated and characterized. To establish the process-property-performance interrelationship, the mechanical, electrical, magnetic, and magnetoelectric properties were exhaustively investigated of DLP processed sample. The mechanical properties were investigated using a standard load frame equipped with a ±1 kN load cell and long travel extensometer. The response of the material to external load will describe the mechanical utility of the material. The electrical properties were characterized based on the hysteretic response collected using the Sawyer-Tower circuit while subjecting the sample to AC electric field. Finally, the magnetoelectric (ME) coupling coefficient was measured using vibrating-sample magnetometer (VSM), inducing an AC magnetization in the sample while measuring the resulting change in voltage. The ME coefficient, defined as the ratio between the resulting electric field to applied magnetic field, is the main performance matrix of the efficacy of the magnetic to electric energy conversion. The results indicate the suitability of DLP additive manufacturing to efficiently produce magnetoelectric composites. Future research will focus of enhancing the properties and further streamline the fabrication process.

The seamless and adaptive interactions between functional devices and their environment (e.g., the human body) are critical for advancing emerging technologies, e.g., wearable devices, consumer electronics, and human-machine interface. The state-of-the-art technologies, however, require a complex integration of heterogeneous components to interface the environmental mechanical stimulus, which is ubiquitous and abundant in the above applications. Moreover, all existing technologies require a power source, which complicates the system design and limits operation schemes.
I will discuss our recent progress in developing self-powered human-integrated nanodevices through the hybrid nanomanufacturing (e.g., 3D printing of self-assembled function materials) of heterostructured nanodevices with hierarchical architectures. This new class of wearable devices are conformable to human skins and can sustainably perform non-invasive functions, e.g., physiological monitoring and gesture recognition, by harvesting the operation power from the human body. This research is expected to have a positive impact and immediate relevance to many societally pervasive areas, e.g., biomedical monitoring, consumer electronics, and intelligent robotics.

Direct ink writing (DIW) is a versatile additive manufacturing technique for patterning a wide variety of materials in three dimensions and is particularly well suited for printing assemblies of multiple functional materials, with electrical, optical, structural or biological properties. Currently, DIW is hindered by a relatively low throughput, which limits the potential applications and suitability for larger-scale manufacturing. To increase the throughput of DIW while retaining its unique advantages, multinozzle DIW printheads have been developed in order to accommodate for the simultaneous deposition of up to 64 filaments. However, this process has been constrained to printing on flat substrates, thereby excluding its use for the functionalization of pre-existing surfaces. To circumvent this shortcoming, we designed an adaptive multinozzle printhead and combined it with non-contact surface profilometry for high throughput printing on non-planar surfaces. The printhead is comprised of 16 nozzles positioned in a linear array and individually actuatable by a set of stepper motors in order to conform its geometry to the underlying substrate topography. We demonstrate that this novel printhead can successfully and rapidly deposit viscoplastic inks onto surfaces with complex profiles. More importantly, we show that this adaptive multinozzle approach can be applied to restore the mechanical integrity of damaged solids via the conformal extrusion of structural elastomeric inks or cover wound regions of human limb models using biological inks.

Biomaterials that closely mimic the complex structural organization of natural tissues hold great promise for regeneration of tissues. Hydrogels have emerged as leading candidates to reproduce native extracellular matrix. In order to provide structures and functions similar to native tissues, controlled porosity and vascular networks are required. However, fabrication techniques to introduce these are still limited. In this study we propose a comparison on fabrication techniques to achieve 3D vascular networks using water based solvents only. Diagnostic methodologies to optimize printing parameters and mechanical characterization are performed on the formulated hydrogel compositions.
With the adequate choice of ingredients and fillers for photocurable formulations, structural and functional properties of the fabricates scaffold can be tailored, opening the path for advanced applications.
Exploring both natural and synthetic hydrogels formulations, fabrication of vascular networks using Digital Light Processing (DLP) is assessed. An analysis of the achieved vascularization morphologies is performed to yield a library of fabrication parameters and hydrogel formulations for future applications.

Polymer additive manufacturing (AM) is a rapidly growing technology and it is transitioning to become an advanced manufacturing technique with the introduction of fibrillar reinforcing materials and the recent developments in large scale AM. While carbon and glass fibers are mainly used as reinforcing phase in polymer composites, as a result of increasing environmental and long-term sustainability concerns, there is an increasing interest in using bio-derived cellulose fibers to reinforce polymers instead. The ability of freeform manufacturing of parts with complex geometries makes large scale AM attractive for various applications; however, different applications require different material properties. One of the interesting application areas for large scale AM is printing of lightweight materials via extrusion foaming. While density is a key property in light weighting, mechanical performance is also important for many lightweight applications. The purpose of this study is to understand the impact of microcellulose fibers (MCF) on foaming behavior and the mechanical properties of additively manufactured parts. MCF-PLA feedstock pellets at varying MCF content (5, 10, 15 and 20wt%) were prepared to understand the impact of cellulose fiber content on density and mechanical properties of the AM biocomposites. Also, the impact of foaming agent content as well as extrusion rate on the AM biocomposites are investigated. Although achieving printed parts with uniform foam structure with the presence of cellulose fibers was challenging, initial results are promising, reaching density values below 0.5g/cm³.

Cell growth and cell adhesion on biocompatible polymers are affected by the surface energy and the interfacial properties. One of the major factors that affect adhesion is contamination by different microorganisms. This contamination and simultaneous growth of microorganisms can affect the quality and quantity of healthy cells that grow on the surface. Polylactic Acid (PLA) is a biodegradable polyester material produced from renewable resources like biomass. Additionally, its biocompatibility has permitted its use in the food packaging and the biomedical industry. Despite its benefits, PLA intrinsically lacks in strength, durability, and adhesiveness. This study focuses on mitigating these problems by incorporating surface modification techniques to increase the surface roughness of PLA. PLA samples were 3D printed and then treated with Corona discharge, a method widely used in a variety of industries because of its safety, low cost, and overall effectiveness. After treatment and surface modification, the material properties of the samples were characterized by using Impedance Spectroscopy, Raman spectroscopy analysis, and Optical Profilometry. The results of these studies showed a significant improvement in the surface properties that promote cell affinity and adhesion.

Ocular surface diseases including conjunctival disorders are multifactorial progressive conditions that can severely affect vision and quality of life. With more than ten million new diagnosis worldwide each year, patients suffering from these severe forms of ocular diseases will often need surgical intervention to regenerate the ocular surface, especially the conjunctiva, to restore vision. With the development of advanced regenerative medicine and stem cell technologies, growing attention over the past decade has turned towards the utilization of stem cell therapy for ocular surface diseases, and stem cell therapies based on conjunctival stem cells (CjSCs) have become a potential solution for treating ocular surface diseases. However, neither an efficient culture of CjSCs nor the development of a minimally invasive ocular surface CjSC transplantation therapy has been reported. The goal of this study was to develop a robust in vitro expansion method for primary rabbit-derived CjSCs and applied digital light processing (DLP)-based bioprinting to produce CjSC-loaded hydrogel micro-scaffolds for injectable delivery. Expansion medium containing small molecule cocktail that performed dual SMAD signaling inhibition (dSMADi) and ROCK signaling inhibition (ROCKi) generated fast dividing and highly homogenous CjSCs for more than 10 passages in feeder-free culture while maintaining the expression of stemness markers (ABCG2, KRT14, P63). With our customized DLP-bioprinting system, the elastic modulus of gelatin methacryloyl (GelMA) hydrogel micro-scaffolds was tuned to enable the 3D culture of CjSCs while supporting their viability, stem cell phenotypes, and the differentiation potential into conjunctival goblet cells. Our bioprinting system also enabled rapid fabrication of 18 cellularized hydrogel micro-scaffolds simultaneously with customizable geometries in less than a minute. These hydrogel micro-scaffolds were well-suited for scalable dynamic suspension culture of CjSCs. The bioprinted hydrogel micro-scaffolds in (100 μm diameter and 100 μm in height) were shear thinning and successfully delivered to the bulbar conjunctival epithelium via minimally invasive subconjunctival injection. This is the first report on the development of bioprinted injectable CjSC-loaded hydrogel micro-scaffolds and the establishment of protocols for robust in vitro expansion of CjSCs. Overall, this work serves as an important framework for understanding the conjunctival stem cell population, conjunctival epithelial biology, as well as the application of CjSCs as a clinically translatable strategy for minimally invasive treatments of severe ocular surface diseases.

The design of new alloys for metal additive manufacturing (AM), or 3D printing, is an essential research field to encourage higher adaption of AM in the industries. Currently, pre-alloyed powders are typically used in metal AM. Pre-alloyed powders are expensive and inflexible in terms of chemical composition. In-situ alloying, which makes use of powder blends, enables high-throughput experimental alloy design and screening. This production approach allows high flexibility in varying the chemical composition.

For bulk sample production, laser powder bed fusion of elemental powder blends was applied to build up titanium-tantalum alloy as proof of concept. Tantalum is an excellent choice for alloying with titanium for biomedical applications due to its' high biocompatibility, corrosion resistance and good mechanical properties. Furthermore, titanium–tantalum alloys are promising materials for such applications because of high strength to modulus ratio. Despite the promising characteristics, they are still not widely used due to the difficulty in alloying titanium with tantalum as a result of their difference in melting point and density. In-situ alloying has been shown to be an effective approach in creating these alloys. The material processed were characterised using optical microscopy, electron backscatter diffraction, scanning electron microscopy, energy-dispersive X-ray spectroscopy, X-ray diffraction, hardness and tensile testing.

The advent of additive manufacturing has enabled many complex, optimized structures to be realized efficiently and reliably, improving the performance of many applications where these architected materials are employed. Nevertheless, many of the materials that are currently compatible with 3D printing exhibit mechanical properties that are far from the desired levels, significantly curtailing the benefits that this technique can provide. To address this, we have developed a method of incorporating graphene nanoplatelets into a commercial PMMA-based photosensitive resin and successfully demonstrated the 3D printing of graphene-polymer composites. The static and dynamic (i.e. viscoelastic and high strain rate) mechanical responses of the printed structures were characterized and found to improve tremendously with as little as 0.02wt% of graphene addition. An important reason for this improvement was that the 3D printing process of graphene-polymer resins involved the Selection of Naturally Aligned Graphene (SNAG), which lined the graphene platelets along the printing axis, reinforcing the stiffness and strength of the material in this direction. The effect of various processing parameters, such as the use of post-print baking and addition of acetone to disperse the graphene nanoplatelets, will be presented in detail as well.

Due to the parallelized nature of digital light processing (DLP), additive manufacturing based on this technique has the unique capability to deliver fine resolution over a large area with reasonable print times. This is advantageous in applications such as tissue bioprinting, where micron-scale complexity persists throughout a tissue which may be tens of centimeters in length. In practice, such high resolution is difficult to achieve due to diffusion of reactive species between light and dark regions of the printing plane. Partially crosslinked macromolecules diffuse away from initiating regions while oxygen and other inhibitors diffuse into them. The combined effect is overpolymerization in dark zones, undercuring in light regions, and a loss of print fidelity. Additionally, gradients within polymerized structures cause mechanical heterogeneity in the final part. Because these processes occur on the individual voxel scale, measurement techniques such as photorheology and infrared spectroscopy are unable to adequately measure them, and new measurement techniques are needed.

In this talk, I will describe an atomic force microscopy (AFM) technique to directly map the generation and diffusion of polymeric species during localized photopolymerization. We use an AFM cantilever with a nanocylinder extending from the tip, and when the cylindrical probe is inserted into a resin, the drag on the cylinder can be used to measure to the local viscosity of the liquid, and in turn, the local extent of reaction. Vibrating the cantilever at one of its resonance frequencies (10⁴ Hz - 10⁷ Hz) affords high temporal resolution. Computational fluid dynamics modeling indicates that the fluid motion induced by the oscillating cantilever is localized within 1 μm of the probe. Employing a custom-built instrument, we can direct arbitrary light patterns to resin in the imaging plane of the AFM to achieve photopolymerization. By translating the tip and light pattern relative to one another, the full spatial evolution of the reaction profile can be measured.

We employed this technique to measure the reaction dynamics during DLP photopolymerization of a thiol-ene resin. The thiol-ene click reaction has some advantages over the photopolymerization of acrylate resins, including reduced shrinkage stress and insensitivity to oxygen inhibition. We find a non-reciprocal dependence of the reaction on light intensity; increasing the light intensity decreases the efficiency of the reaction, likely by increasing the bi-radical termination rate. Thus, at constant light dose, lower light intensities lead to increased conversion in the reaction volume, even accounting for increased time for diffusion. This disproves a common assumption in stereolithography, namely that gelation depends solely on the light dose, often referred to as the “critical dose.”

Mapping the spatial evolution of the polymer conversion profile also allows us to easily identify the diffusion length scales during the reaction, and we measure polymer accumulation 20 μm away from the illuminated region within one second of light exposure, identifying the relevant length scale of our process. We then created an open ring test structure (inner diameter = 15 μm, outer diameter = 45 μm) based on this finding to determine the effect that diffusion plays on the resolution of a printed part. Consistent with our measurements of the reaction profile, we observe significant overpolymerization in the ring center, leading to gelation and loss of fidelity to the projected mask. This new technique provides adequate spatial (1 μm) and temporal (<1 ms) resolution for modern stereolithography applications, and it can be used to predict the ultimate printing resolution and rapidly determine the optimal exposure conditions for a given resin.

3D printing can be used to create prototypes and devices from polymeric materials which have appended the design functionality for new materials including uses in biomedical devices enabling their rapid development. While 3D printed polymers can be further classified into thermoplastics, thermosets, and
elastomers based on their thermo-mechanical properties – new opportunities for multi-materials and
composites are possible. The processability and functionality of thermosets and elastomers make it a
challenge to employ using most 3D printing methods for polymer additive manufacturing. The transition to a
final phase or cross-linked structure results in new properties in combination with the processing method. However, 4D printing allows the design of new materials and applications based on
integrating the chemistry of conversion with the printing mode and final stimuli-responsive property on the printed part. In this talk, we will demonstrate the 4D fabrication of multi-materials including thermosets and thermoset elastomers with concept objects and elastomeric properties unlock with stimuli-response. This is based on the use of extruded viscous solutions. The result is an extrudable precursor and nancomposite elastomers which can be printed via viscous extrusion printing (VEP) or viscous solution printing (VSP) and then converted to an elastomeric actuating material with very high cyclic compressibility or a shape memory thermoset. Other works based on the use of SLA, SLS, FDM, towards high strength epoxy, silicones and nanocomposite materials will be discussed.

The convergence of three-dimensional (3D) printing of cell seeded bioinks and regenerative medicine offers promise in the biofabrication of tissue building blocks with tailored structural, biological and mechanical properties. However, current 3D bioprinting approaches are limited in their ability to print fibrous collagen bioinks at different length scales that replicate the complex hierarchical architecture of native tissues. Collagen-based hydrogel bioinks are restricted by their narrow printability range, where protein structure, seeded cell viability, and bioactivity of incorporated biomolecules all need to be maintained within physiological boundaries. Herein, a unique approach to biofabricate tissues has been developed to overcome these challenges. Automated gel aspiration-ejection (GAE) leverages the properties of dense collagen gels as tissue models and templates for regenerative medicine. Automated-GAE fabricates highly defined mini-tissue building blocks of various dimensions and shapes, e.g., cylindrical, quadrangular and tubular as well as with tunable microstructural and mechanical properties that modulate seeded cellular responses.
Acellular and cell seeded cylindrical highly hydrated precursor gels were initially cast in different volumes. Automated-GAE was carried out using a uniquely configured instrument equipped with multiple syringe pumps connected to various densification needles to generate dense collagen gel bioinks of different sizes and microstructures. Through the creation of a pressure differential, GAE aspirates the precursor highly hydrated gels into the needles, expelling their unbound fluid and simultaneously inducing their compaction and meso-scale anisotropy. By subsequent reversal of the pressure, dense collagen gel bioinks can be controllably ejected that can be cylindrical, quadrangular or tubular in shape. The incorporation of bioactive materials can also facilitate the production of rapidly mineralizable tissue blocks.
A simple mathematical relationship defining the bioink compaction factor by relating changes between initial highly hydrated hydrogel precursor and final dense gel dimensions was found to be surprisingly highly effective in predicting the initial and temporal properties of the bioinks in culture. Bioink collagen fibrillar density and alignment, tissue building block mechanical properties, as well as seeded cell density, morphology, viability and proliferation were all quantitatively correlated with fabrication compaction factor. Furthermore, by controlling initial bioink parameters, seeded cellular function, in terms of differentiation and extent of matrix remodelling ability were modulated. To this end, automated-GAE will potentially also enable a fourth dimension to biofabrication, where cell-cell communications and cell-extracellular matrix interactions as a function of time in culture can be predicted and modelled.
Therefore, by initially using naturally derived reconstituted collagen hydrogels incorporating cells, and negating the need to directly extrude pre-polymerized collagen molecules, the automated-GAE fabricated mini tissue blocks can be tailored to mimic protein fibril density and alignment of target native tissues, as well as cell loading, density and orientation according to the intended use. Furthermore, the mini tissue blocks can be combined to generate more intricate structures resembling those found in native tissues. Thus, a pre-defined microenvironment can be designed and tuned along with controlling cellular function to meet specific structural requirements of both, mineralized and soft tissues. This breakthrough development will also potentially enable the rapid 3D printing of tissues with varying architectures based on a platform bioink system.

Hydroxyapatite/collagen nanocomposite (HAp/Col) is a material mimicking bone nanostructure and chemical composition and the world first material reported as completely incorporated into bone remodeling process and substituted with new bone. Porous HAp/Col has been already applied in Japan as a bone void filler and is also considered as a scaffold for bone tissue regenerative medicine. The HAp/Col contains collagen molecules; thus the porous body is prepared by freeze casting and pore structure is only controlled by ice crystals formation in its suspension. Since pore structure influences on cell behaviors including activation of various functions, control of pore structure is one topic for the HAp/Col functionalization. Extrusion type 3D printing is a good technique to fabricate designed pore structure for collagen containing materials. Alginate-based and silane coupling agent (SCA)-based HAp/Col pastes were prepared for 3D printing. The alginate-based HAp/Col paste was immediately hardened to viscoelastic body by extruding into Ca²⁺ containing solution and can be applied to fabricate cell-scaffold construct. The SCA-based HAp/Col paste maintains its shape even if it extrudes directly into water and hardened in 30 min.
The SCA-based HAp/Col pastes were directly injected into bone hole of SD-Rat and remained the injected site by hardened. They were started to remodeled in 7 days without any bad symptoms. The SCA-based HAp/Col paste injected directly into holes of porcine tibia completely substituted with new bone in 3 months without any bad symptoms. The HAp/Col pastes are good candidate for fabrication of cell-scaffold constructs in bone and maybe other tissues due to their bioresorbable and viscoelastic nature.

Personalized medical device fabricated by additive manufacturing is a booming research field for it enables innovative solutions to cater for patients’ specific needs. Seemingly, the predominant limitations to scale up in the medical industry arise from the suboptimal biomechanical properties and challenge in boosting the productivity. In this study, we introduce a hybrid laser sintering-heat treatment approach to address this problem. We first examine the microstructure of additive manufactured components under various sintering and heat treatment parameters, and thereby reveal the optimization in terms of material and biomechanical properties, efficacy of bio-functions, and efficiency in cost and time of manufacturing. In addition, the biocompatibility of the 3D printed alloy is validated via haemolysis assay, cytotoxicity and implantation tests. We further present a pathway from proof-of-concept to clinical application with our state-of-the-art examples. Our work will shed lights on the industrialization of additive manufacturing in medical device industry.

Hydrogels are promising material candidates in diverse applications including drug delivery, tissue engineering and soft robotics, owing to their excellent properties of biocompatibility, wetness, softness, deformability and responsiveness [1]. However, the intrinsic activeness of hydrogels for autonomous locomotion is not achieved yet. The existing activeness of hydrogels are realized by capping active nanoparticles like MnO₂ and TiO₂, which undergo relative catalytic reactions to perform tasks for environmental applications [2,3]. Furthermore, we fabricated versatile printable active hydrogels which possesses the ability to move on water surface. Utilizing the surface tension, the homogeneous active hydrogels propel themselves and show well-controlled and intelligent locomotion on water surface. Incorporated with stimuli-responsive groups, the active hydrogels are able to change their geometries with time under relative stimulus, which adds the dimension of time (4^th dimension) to the 3D-printable active hydrogels. The processing technique of these active hydrogels are fully compatible to existing photo-curing 3D-printing equipment. These active hydrogels provide new opportunities for 3D-printing technology to be applied in the field of miniaturized robots.

Reference:
[1] Xinyi Lin, et al. Research 1-15 (2020).
[2] Xinyi Lin, et al. Adv. Mater. Technol. 5 1-10 (2020).
[3] Jinrun Liu, et al. Environ. Sci.: Nano, 7 656-664 (2020).

3D printing is ideal for prototyping yet its low productivity prevents it from being further applied in large-scale manufacture. Some innovative techniques have been developed to break the limitation of printing speed, however, sophisticated facilities or costly consumables are required, which still substantially restricts the economic efficiency. Here we report that a most common stereolithographic 3D printing facility can achieve a very high printing speed (400 mm/h) using a green and inexpensive hydrogel as an anti-adhesion interface (release films). The anti-adhesion mechanism relies on large deformation of the soft hydrogel, which is distinct from that of other techniques. Adhesive force of the solid-solid interface during the layer-by-layer photo curing can be largely reduced according to such mechanism. The technique shows an excellent printing stability even for fabricating large continuous solid structures, which is challenging for other rapid 3D printing techniques.

A commercialized fluorescence-linked immunosorbent assay (FLISA) is a quantitative technique for detecting bio-chemicals through antigen-antibody binding reaction using well-plate platform. As the manufacturing technology of microfluidic systems has improved, the FLISA is implemented onto the microfluidic disk platform which shows detection of trace bio-chemicals with high resolution. It is possible to reduce time from the reagent loading to the detection step within an hour, in addition to benefits of reducing the amount of reagents to 1/10. Since the high density antibodies on the surface of microbeads, the potential is increased for immobilization of bio-molecules. The incubation process requires not only the binding the antigen-antibody, but also the several steps for binding fluorogenic substrates to target protein. And the unbound reagents should be removed by a washing process. The FLISA protocol in the microfluidic platform, it is necessary to properly execute of liquid reagents movement in each process of FLISA in order to perform the binding reaction sufficiently. The precise control of reagent liquids is required such as fluidic isolation of the incubation chamber and repetitive angular rotation control of disk. We suggest a novel microfluidic disk including a 3D incubation chamber using multi-material to low cost which is fabricated to simple mechanical assembly. VEGF detection is performed sequentially one-step using a 3D microfluidic disk within an hour. On the 3D microfluidic disk, the microbead-based FLISA protocol is operated from the incubation process to the detection process in hand-free without additional components to liquid control. For the incubation process, the microbead movement is controlled through the centrifugal force by disk rotation and gravitational force by bead sedimentation on a slope of chamber.

Several in vitro neural network models have been developed to mimic the reconstruction and interconnection of neural networks and to study brain function and related diseases. Such in vitro neural networks require predefined neural connections at the target location and the measurement of neural activity using a multi-electrode array (MEA). The predefined neural connections sometimes limit the adaptation of the user to the change in the neural connection patterns. Here we report a three-dimensional (3D) magnetically actuated microrobot fabricated by 3D laser lithography based on two-photon polymerization for selective neurite alignment and neuronal connections. Using confocal immunofluorescence imaging, the aligned neurite outgrowth and synaptic connections in the neural network were measured. The microrobot, in which the rat's primary hippocampal cells were cultivated, was transmitted between two neural clusters by a magnetic field and then structurally and functionally connected to a neural network that can transmit neural activity signals. Neuronal activities were measured with a complementary metal-oxide-semiconductor-based MEA system to monitor the propagation of extracellular axonal signals from the neural clusters. The proposed microrobot shows the potential for in vitro neural experiments to understand how neurons communicate in the neural network by actively connecting neural clusters.

Microneedles are a transdermal drug delivery system with length dimensions of several hundred micrometers. To be considered in biomedical applications, the biocompatibility and safety of materials are critical factors. The use of protein-based biomaterials such as silk fibroin(SF) can clearly reduce immunological risks resulting from accidental breakage of microneedles during injection into the skin. In the experiment, one-step 3D structuring of SF-based microneedles is challenged using riboflavin as an enzymatic photoinitiator. Digital light processing (DLP) 3D printing is adopted because it is advantageous for direct 3D construction of molecules dissolved in aqueous solution at relatively low concentration through photocrosslinking. Oblique and sharp microneedles are printed using an anti-aliasing strategy and shrinkage of the hydrogel in a dehydration process. The SF microneedle is strong and showed no severe damage to the structure upon application of ~300 mN of compressive stress. A flexible SF microneedle array pad is fabricated and subsequently utilized for delivery of fluorescence dye molecules into pig skin.

4D printing gives 3D printed structures shape morphing properties when exposed to external stimuli. Many such materials have been reported using hydrogels, liquid crystals and shape memory polymers. However, these materials generally lack the mechanical strength and stiffness required for common applications in the robotics and aerospace industries. In this work, we employ direct-ink-writing to fabricate stiff fibre-reinforced epoxy composites which exhibit temperature controlled reversible morphing. The composite is printed flat initially and the morphing behaviour is activated by heating the material to a temperature above its glass transition temperature. The reversible morphing is enabled by the microstructuring of the printed material arising from the shear-induced fibre alignment during printing. This local alignment controls local anisotropy in the material properties that drives deformation upon heating.

The experimental results, along with finite elemental analysis reveal that by varying the layer print orientation and activation temperature of the prints, the resultant shape and curvature of the morphed material can be controlled precisely as desired. Using an appropriate print design, structures which exhibit high temperature bistability could be achieved, which allows reversible morphing into multiple stable shapes in stiff materials. Furthermore, the ink composition can be tuned to generate shape-dependent functional properties, such as electrical conductivity. Therefore, our method has the potential to fabricate functional composites with tailored made morphing behaviours for applications such as actuators and sensors.

High performance microprobes for combined sensing of glucose and choline were fabricated using microcontact printing (μCP) to transfer choline oxidase (ChOx) and glucose oxidase (GOx) onto targeted sites on implantable microelectrode arrays (MEAs). Many neuroscience studies require the monitoring of multiple species, including metabolites and neurotransmitters, with high spatiotemporal resolution in vivo. Choline is of interest as a surrogate for the neurotransmitter, acetylcholine, which is turned over rapidly by acetylcholinesterase in the brain. Most electroenzymatic sensing sites on MEAs for neuroscience applications have been created by manual enzyme deposition, however this approach becomes problematic when the array feature size is less than or equal to ∼100 μm. The μCP process used here relies on use of soft lithography to create features on a polydimethylsiloxane (PDMS) microstamp that correspond to the dimensions and array locations of targeted, microscale sites on a MEA. Precise alignment of the stamp with the MEA is also required to transfer enzyme only onto the specified microelectrode(s). The dual sensor fabrication process began with polyphenylenediamine (PPD) electrodeposition on all Pt microelectrodes to block common interferents (e.g., ascorbic acid and dopamine) found in brain extracellular fluid. Next, a chitosan film was electrodeposited to serve as an adhesive layer. The two enzymes, ChOx and GOx, were transferred onto different microelectrodes of 2 × 2 arrays using two different PDMS stamps and a microscope for stamp alignment. Using constant potential amperometry, the combined sensing microprobe was confirmed to have high sensitivity for both choline and glucose (286 and 117 μA mM^-1 cm^-2, respectively) accompanied by low detection limits (1 and 3 μM, respectively) and rapid response times (≤2 s). The performance of the dual sensing microprobe was validated in the dorsal striatum of anesthetized rats. This work demonstrates the use of μCP for facile creation of multi-analyte sensing microprobes by targeted deposition of enzymes onto preselected sites of a microelectrode array. Such technology will enable neuroscientists to monitor multiple species at the same location in the brain and to integrate chemical sensing sites with microelectrodes for electrophysiological recordings so as to better correlate neurochemical signaling with neuronal activity.

We synthesize 3D-printable inks containing live diatoms and diatom nutrients. The glass skeletons of the diatoms form the basis for structures of high complexity and with many levels of hierarchy reaching from the nanometer scale to the centimeter scale. The diatoms survive the printing process and keep reproducing in the printed structures. We have succeeded implementing this method for several types of diatoms featuring different morphologies and sizes. The printed structures can be converted into several types of material, such as composites with diatoms as the reinforcing agent, or into materials entirely consisting of glass. Due to their nano-structured morphology, the highly heat resistant all-glass materials exhibit significantly enhanced mechanical properties compared to bulk glass, while being much lighter. Based on home-grown biological feedstock, we consider this a promising approach for the scalable production of sustainable performance materials with a negative carbon footprint and widely tunable properties and functionality.

Tissue engineering offers a promising approach to treat difficult problems in human body tissue repair. It involves using live cells to form implantable devices for body tissue regeneration. In scaffold-based tissue engineering, a porous scaffold provides a microenvironment for cells to adhere, proliferate and differentiate and a structural framework for new tissue formation. Most human body tissues are complex and their regeneration requires structurally complex scaffolds that resemble tissue structures and can provide biochemical cues such as growth factors (GFs). Incorporating GFs and even live cells in scaffolds can greatly facilitate tissue regeneration. 3D printing has many advantages in scaffold fabrication, such as control of pore shape, size, porosity, etc. Furthermore, 3D printing can make complex tissue engineering scaffolds, including multilayered scaffolds with different layer characteristics for regenerating body tissues that exhibit multilayered structures such as osteochondral tissue. Therefore, like in other industries, 3D printing has already made a high impact in the tissue engineering field, with numerous researchers around the world using 3D printing technologies to produce various tissue engineering products. 4D printing emerged in 2013 and immediately attracted world’s attention. 4D printing uses 3D printing technologies to produce shape-morphing objects. Such objects can meet the demanding requirements in particular applications. The concept of 4D printing has been evolving and one current popular definition of 4D printing is that the shape, property and functionality of a 3D printed object can change with time in a predefined design. 4D printing relies on advances in 3D printing technologies, smart materials, and smart designs. It has become an actively pursued subject in both academia of different disciplines and industry, including tissue engineering. We have conceptualized 5D printing and are applying it in tissue engineering. 5D printing produces shape-morphing and information-embedded structures, and the information, which is the 5th dimension in 5D printed structures, is delivered in situ during applications of these structures. More importantly, with 5D printed structures, the delivered information affects the surrounding environment (or 5D printed structures) and guide changes in the environment (or 5D printed structures). For 5D printing in tissue engineering, the embedded information can be biomolecules such as GFs. Using 5D printing, shape-morphing, mesenchymal stem cell (MSC)-containing and GF-delivering multilayered complex scaffolds may be constructed for gastrointestinal (GI) tissue engineering. Under GFs’ guidance, MSCs will differentiate into different types of cells in the cell-scaffold constructs for GI tissue regeneration. This talk will present our work in 3D/4D/5D printing of scaffolds for body tissue regeneration. It will focus on the design and construction of complex tissue engineering scaffolds.

3D printing has greatly improved our ability to create complex tissue engineering scaffolds with high precision. Biomaterials, growth factors or even living cells are accurately deposited layer-by-layer in 3D printing to construct the scaffolds. Hydrogels have been commonly used biomaterials in 3D printing of scaffolds owing to their various advantages. However, most hydrogels, particularly natural hydrogels, have poor mechanical properties, which severely limit their tissue engineering applications. It has been shown that adding polymer nanofibers into hydrogels can lead to improved mechanical properties that can be comparable to the body tissues. Growth factors (GFs) in the body can promote wound healing, cell growth, proliferation and differentiation. Incorporating GFs into scaffolds may accelerate tissue regeneration. Fibroblast growth factor (FGF) is generally used in the regeneration of tissues such as cornea and skin. In this study, nanofiber reinforced scaffolds with the controlled release of GF were investigated with 3D printing of PLGA nanofiber (PLGAf) reinforced alginate hydrogel containing FGF. To control the FGF release, two strategies were adopted: (1) FGF was directly included in the biomaterial (PLGAf and alginate hydrogel mixture) for 3D printing, (2) FGF was firstly encapsulated in PLGA nanofibers via emulsion electrospinning and the nanofibers were then dispersed in alginate hydrogel for 3D printing. Thus two types of inks were made for 3D printing: FGF/alginate/PLGAf ink, and alginate/PLGAf-FGF ink. They were used in an extrusion-based 3D bioprinter to fabricate reinforced scaffolds according to the CAD design. The experiments showed that the addition of PLGAf into alginate hydrogel greatly improved its viscosity and mechanical strength. In vitro release studies were conducted for scaffolds 3D printed from the two type of inks. Release results showed that both types of scaffolds exhibited two-stage release profiles: an initial fast release period in the first 2 days, and the subsequent slower and sustained release period. In addition, alginate/PLGAf-FGF scaffolds displayed a slower release than FGF/alginate/PLGAf scaffolds, which was mainly due to the different incorporation site of FGF within the scaffolds. It can be concluded that the GF release behaviour from 3D printed scaffolds can be controlled through choosing the loading site in scaffolds for GFs, which enables designing personalized GF delivery specific for the targeted tissue engineering application.

Three-dimensional (3D) printing offers the unparalleled capability to create medical devices with complex architectures matched to the patient’s anatomy. However, most of the currently used polymers require high processing temperatures and pressures and/or a combination with leachable additives like initiators, crosslinkers, plasticizers, and solvents to enable extrusion-based direct-write 3D printing (EDP). Such conditions may raise safety concerns for the final medical product. Therefore, it is desirable to develop polymers that could be printed at ambient conditions and without additives. To develop such polymers, it is necessary to systematically understand the relationship between polymer molecular structure, rheology, and 3D printability. Herein, we present a library of novel polyesters with modular functionalities, which can be used to create customized 3D constructs using EDP. The polyesters with various pendant functional groups were synthesized and their physical properties, rheology, and 3D printability were analyzed.
Methods: The polyesters were synthesized at room temperature using carbodiimide-mediated polymerization of pendant functionalized diols and succinic acid, and characterized using ¹H-NMR and variable temperature-FTIR. Polymer viscosity as a function of shear rate was measured under steady state shear flow. Small amplitude oscillatory shear experiments were done to plot master curves of viscoelastic properties for each polyester over a wide frequency range using the time-temperature superposition (TTS) principle. The 3D printability of the polyesters was assessed based on parameters such as the ability to extrude polymer as continuous filaments through a narrow nozzle at a consistent flow rate, retain the printed shape, form bridge-spanning filaments without significant sagging or collapse, and form multilayer constructs.
Results: Data demonstrated that the synthesized polyesters exhibit properties that are desirable for the extrusion-based 3D printing. ¹H-NMR confirmed the presence of the appropriate functional groups for each synthesized polyester suggesting successful synthesis. All synthesized polyesters exhibit shear thinning behavior at EDP relevant shear rates and respective 3D printing temperatures, which is important for solvent-free printing. The tangent delta peak observed at temperatures higher than polymer T_g indicated the presence of a “secondary network” and supramolecular interactions. Variable temperature-FTIR supported the presence of supramolecular interactions, such as H-bonding. The presence of “secondary network” along with supramolecular interactions appear to facilitate the retention of printed shape and improve the overall quality of the 3D printed constructs. Based on observed correlations between 3D printability and the rheology of each polyester a “3D printability window” is proposed. The insights derived from this study can be used to inform the design of new biodegradable polymers for extrusion-based direct-write 3D printing for biomedical applications.

The ability to precisely control the localization of enzymes on a surface is critical for several applications including biosensing, nano-bioreactors and single molecule studies. Despite recent advances, fabrication of enzyme patterns with resolution at the single enzyme level is limited by the lack of lithography methods that combine high resolution, compatibility with soft, polymeric structures, ease of fabrication and high throughput. Here, a method to generate enzyme nanopatterns on a polymer surface is demonstrated using thermochemical scanning probe lithography (tc-SPL) and the enzyme Thermolysin as a model system. Electrostatic immobilization of negatively charged sulfonated enzymes occurs selectively at positively charged amine nanopatterns produced by thermal deprotection of amines along the side-chain of a methacrylate-based copolymer film via tc-SPL. This process occurs simultaneously with local thermal quasi-3D topographical patterning of the same polymer, offering lateral sub-10 nm resolution and vertical 1 nm resolution, as well as high throughput (5.2 x 10⁴ mm²/h). The obtained patterns with single enzyme resolution are characterized by atomic force microscopy (AFM) and fluorescent microscopy. The enzyme density, the surface passivation and the quasi 3D arbitrary geometry of these patterned pockets are directly controlled in a single step, without the need of markers or masks. Other unique features of this patterning approach include the combined single-enzyme resolution over mm² areas and the possibility of fabricating enzymes gradients at the nanoscale.

With the development of stem cells, in vitro tissue engineering has become a promising technology for research, clinical testing, and medical treatment. One consistent focus of research has been the high-precision patterning of cells, necessary because material surface properties, environment, and organization of cells in two- and three-dimensions all significantly impact the growth and development of tissue. While many different cell patterning techniques have been explored for cell patterning, the integration of organic electronic processes has been previously overlooked. In this presentation, we demonstrate a process whereby organic semiconductors are used to pattern cells and discuss the unique advantages of such a process. Using vacuum thermal evaporation (VTE) and organic vapor jet printing (OVJP), two organic electronic materials are precisely deposited in the vapor-phase as adhesion points on a biocompatible poly(p-xylylene) surface. The small molecular weight organics prevent the subsequent growth of anti-fouling polyethylene glycol methacrylate (PEGMA) polymer brushes, rendering the background areas of the substrate protein and cell resistant. Fibronectin then attaches to the deposited organic semiconductor regions, followed by the selective adhesion of fibroblasts. For VTE, the adhesion regions consist of hexagons with side-lengths of 25 µm, while OVJP is used to form patterns varying in width from several hundred micrometers to about 10 μm. At smaller widths, cells show considerable alignment to the OVJP-formed patterns, moving to random orientation at a threshold width of about 110 µm. The surface properties of the different organic semiconductor materials also prove important, demonstrating different patterning tendencies. The cell-patterning process presented here diverges from other techniques in i) the usage of organic small-molecule semiconductors as adhesion regions, ii) the formation of adhesion via vapor-phase deposition. Some of the many advantages include the vast number of evaporable organic semiconductor materials, the ability to deposit onto a fragile scaffold without contact, and the potential for high-throughput manufacturing. Therefore, this process is a useful development in cell patterning with many potential applications.

We successfully developed a complete process chain of production lines for foil-based bioanalytical lab-on-chip devices. The process is based on high-throughput structuring and lamination using roll-to-roll (R2R) UV micro or nano-imprinting on several hundred meters-long flexible, polymeric foils. Bio-functionalization of the imprinted structures is achieved via R2R microarray printing. In our pilot line, the polymer foil is roto-gravure coated using a photopolymer with tuneable properties, which is then structured via R2R UV nanoimprint lithography (NIL).^[1] The technology allows producing various kinds of fluidic structures; such as, capillary force-driven fluidic channels, reservoirs or pumps as well as production of optical structures with different geometries for effective guiding of light for in-vitro diagnostic (IVD) products. Following R2R UV NIL imprinting, bio-functionalization is achieved using a custom-made, semi-automated R2R microarray spotting unit. For specific bio-detections via IVD chips, various probe DNAs or proteins can be printed at different sections of biochips. With layout design flexibility and rapid prototyping possibilities, up to 7500 biochips per 100 meters are produced via our technology. The feasibility of the approach for massively parallel production of lab-on-a-foil products is demonstrated based on a model application of in-vitro multiplexed DNA testing for markers of a methicillin resistant pathogen.^[2] The manufactured foil chips show similar detection performance as commercial injection-moulded chips, thus demonstrating that our lab-on-a-foil technology is a potential replacement for the commercially available, disposable chips for IVD applications.
REFERENCES
[1] M. Leitgeb et al., ACS Nano, 10, (2016) 4926-4941.
[2] P. Toren et al., Lab on a Chip, 20, (2020) 4106-4117, featured as inner front cover.
ACKNOWLEDGEMENT
This research was supported by NextGenMicrofluidics project (www.nextgenmicrofluidics.eu) Horizon2020 European Union Research and Innovation Programme with grant agreement n^o 862092.

With vital functions like talking, chewing, and swallowing, an optimized implant should replace a diseased temporomandibular joint (TMJ). Moreover, it is unique as the only bilateral joint in the human body acting as one unit, its movements are a combination of rotations and translations, and it is the most regularly used joint on a daily basis, suggesting a broad range of highly customized TMJ prostheses. In reality, the vast majority of available options contradict these prerequisites, because they are mainly focused on shape reconstruction, rather than functional reconstruction, e.g., no features are foreseen for laterotrusive movements (which are necessary for proper grinding of food) and the implants are not adapted to allow contralateral movements. This leads to partial improvement in functionality (an increase of maximum mouth opening of ±1 cm), but there is room for optimization.

During this research, the entire concept was revised based on all prerequisites and new advances in technology and surgery. Based on experience and various simulations, a thorough analysis was conducted to determine the optimal building blocks for this concept; crosslinked ultrahigh molecular weight polyethylene has shown its excellence in various medical joints, while biocompatible Ti6Al4V (Grade 23, ELI) allows for high customization when using three-dimensional printing (Selective Laser Melting). Combining these raw materials with biofunctionalization (e.g., surface treatments and lattice structures) and a promising HadSat^®-coating has led to excellent wear properties and long-term in vivo results (simulated in a sheep study).

The importance of function-reconstruction translated into specific design parameters; the bilateral nature of the TMJ’s caused the prosthetic side to greatly influence the healthy side (and vice versa) and, with that, all mandibular movements (e.g., opening, closing, protrusive, and lateral movements). To consider these challenges during the patient-specific design of the implant, it was reverse-engineered. Starting from the desired mandibular movements for a specific patient, the topology of the custom-made implant was derived. These desired mandibular movements were based on the pre-diseased anatomy of the patient, using a statistical shape model. This methodology allowed the designers to incorporate important anatomical features (such as condylar path angulation and Bennet movements), and to achieve the optimized patient-specific design.

By incorporating several additional distinctive features, the “bionic” prosthesis was developed. Several lattice structures (scaffolds) were used in the design, to allow for bone ingrowth. This osseointegration served as a secondary fixation method and, together with a lateral movement-restricting lip, reduced the number of necessary screws and thus surgical time. Furthermore, by implementing a scaffold into the condylar neck, the pterygoideus lateralis muscle could be reattached, which helped the patient during lateral movements. This unique feature, together with the abovementioned materials and patient-specific design, next to the sheep experiments, resulted in extensive simulation using finite element analyses to ensure both patient safety and long-term results.

By incorporating all of the abovementioned features into a novel TMJ prosthesis concept, a major advance in function-reconstructive temporomandibular joint replacement was achieved. Early in vivo results (1 year after surgery) showed promising outcomes, involving both high increases in mandibular movements and decreases in pain scores.

Additive Manufacturing has increased our ability to fabricate complex shapes and multi-material structures. Epoxy is excellent as the base for structural composite materials. Furthermore, carbon nanotube (CNT) is an outstanding filler due to its unique properties and functionalities. Here, conductive epoxy-nanoclay-CNT nanocomposite structures were fabricated by direct-write 3D printing. In this process, 3D-printable composite inks were synthesized by incorporation of nanoclay and different concentrations of CNTs – 0.25, 0.5, and 1 vol%, 0.43, 0.86, and 1.7 wt% – in epoxy. CNTs were found to significantly improve the electrical and mechanical properties. Rheological characterization of the inks revealed a shear-thinning behavior for all the nanocomposite inks and an increase in the complex viscosity, storage, and loss moduli with the incorporation of CNTs. The CNT concentration of 0.5 vol% was found to be the optimum condition for enhancement of mechanical properties; an average increase of 61, 59, and 31% was measured for flexural strength, flexural modulus, and tensile strength, respectively, compared to the 3D printed epoxy-nanoclay nanocomposite structures. The electrical conductivity of 2.4 x 10^-8 and 2.2 x 10^-6 S/cm was measured for the nanocomposites containing 0.5 and 1 vol% CNTs, respectively. Multi-scale characterization of the morphology revealed partial alignment of CNTs in the direction of printing, CNT pull-out and breakage at the fracture surfaces, and nano-scale interactions of the constituents, all of which contribute to the superiority of the nanocomposite with CNTs. The findings show the promise of this ink material and printing method for various applications such as aerospace structures and electronics.

This work presents a MXene polymer composite directly 3D printed via binder jet technique. We introduce a strategy to print poly(vinyl alcohol) (PVOH) composite with optimized MXene ink. By ejecting highly conductive MXene particles onto a PVOH matrix, the resulting sample is able to achieve semi-conducting behaviour with the potential for strain sensing and energy storage. The printed component can be used as a strain sensor capable of sensing tensile strains between 11 to 250%. The component could also be used as electrode material in a sandwich-structured pseudo-capacitor with an energy density of 36.4mJcm^3 and a power density of 2.8mWcm^-3. This study demonstrates that binder jet printing has the potential to directly fabricate polymer composite materials end different end applications.

In order to briefly explain why this submission will appeal to broad and interdisciplinary researchers in both 3D manufacturing and 2D materials, it should be noted that although binder jet printing is widely adopted in industrial setting to print ceramic and metallic components. However, it has only been proven very recently as a viable method to print polymer composite materials. The submitted manuscript not only offers a guideline toward ink optimization for binder jet printing with polymeric material, but also for the first time demonstrates different end applications of printed components. This work is believed to be highly capable of providing the readers in the field of 3D printing with a framework on the modification of particle morphology and ink composition in order to further explore the potential limit of binder jet printing of polymeric composite.

Organ phantoms have been recognized as a testing platform in medicine and have become more realistic as the 3D printing technology has advanced. Typical 3D printers can achieve resolutions of tens of microns and offer a number of printing materials [1]. This allows detailed anatomical features to be resolved by choosing different materials [2,3]. While most printing materials do not have properties that reflect those of tissues, 3D printing with biocompatible hydrogels show tissue-like properties [4]. However, most of the work to date has focused on reproducing structural details resemblance, which is mainly of interest for visual comparison.
Here, we present high-fidelity organ phantoms of the full urinary tract that can also be used to simulate and evaluate medical procedures, including surgery. We used 3D-printed molds to build the phantom. The 3D-printed molds were designed with computer-aided design (CAD) and are based on high-resolution X-Ray images. The casting method allows the use of materials that match the mechanical, optical, and imaging contrast properties of biological tissues.
Our phantom has been validated for the simulation and evaluation of many medical procedures, such as lithotripsy [5], cystoscopy [6], and the transurethral resection of the prostate [7]. A major advantage of the phantom is that the surgery can be quantitatively evaluated using multi-modality medical imaging to an extent that cannot be achieved in a real surgery. Both user-feedback as well as the physical parameters of time, accuracy, geometrical centricity, and surface smoothness of the surgical resection can be quantitatively evaluated. These phantom are expected to be important tools for medical training, surgical planning as well as the development of new medical instruments.

[1] V. Filippou et al., Recent advances on the development of phantoms using 3D printing for imaging with CT, MRI, PET, SPECT, and ultrasound, Med Phys 45, e740 - e760 (2018)
[2] F. Adams et al., Soft 3D-Printed Phantom of the Human Kidney with Collecting System. Ann Biomed Eng 45, 963–972 (2017)
[3] K. Qiu et al., 3D Printed Organ Models for Surgical Applications, Annu Rev Anal Chem 11, 287–306 (2018)
[4] J. Li et al., 3D printing of hydrogels: Rational design strategies and emerging biomedical applications, Mater. Sci. Eng. R Rep, 140, 100543 (2020)
[5] D. Li et al., Soft Phantom for the Training of Renal Calculi Diagnostics and Lithotripsy, 41th IEEE EMBC, 19109241 (2019)
[6] E. Choi et al., Soft Urinary Bladder Phantom for Endoscopic Training, submitted
[7] E. Choi et al., A High-Fidelity Phantom for the Simulation and Quantitative Evaluation of Transurethral Resection of the Prostate. Ann Biomed Eng 48, 437–446 (2020)

The integration of electronics with the human body, such as wireless sensors and a drug delivery system can enable a personalized digital-based diagnosis and treatment strategies. Nevertheless, surgically placed medical implants are associated with eliciting foreign body immune responses and can serve as a nidus for infection. Recent advances in 3D printing have enabled the creation of novel 3D architecture and devices with an unprecedented level of complexity, properties, and functionalities. The development of a multi-scale, multi-material 3D printing approach can overcome the geometrical, mechanical and material dichotomies between conventional manufacturing technologies and a broad range of three-dimensional systems. Here, we discuss the creation of 3D printed gastric resident electronics (GRE) that can circumvent the potential complications associated with surgically placed medical implants by leveraging the significant space and immune-tolerant of the stomach. We propose biomedical applications with the further advancement of this system that can be potentially realized with a multiscale 3D printing process that synergistically integrates functional nanomaterials.

3D bioprinting is an enabling tool for regenerative medicine and drug discovery. Despite promising advancements 3D bioprinting technology, a need remains for reproducible commercially available ready-to-use bioinks to replicate 3D tissue microenvironments. To address this, we have developed a platform of bioink formulations, TissueFab^TM, that are compatible with diverse cell types, bioprinting platforms and have high batch-to-batch consistency. TissueFab^TM is a family of general purpose and tissue specific ready-to-use bioinks based on natural proteins and polysaccharides and synthetic polymers. These bioinks are designed for optimal viscosity and mechanical properties for high resolution bioprinting and to maintain high cell viability (>80%) in a wide range of curing wavelengths. Printability of our bioinks are assessed on multiple microextrusion based commercial bioprinters thus demonstrating the printer agnostic characteristic of the bioinks. Bioink formulations were validated for high cell viability, proliferation and metabolic activity using C2C12 mouse myoblast cells, human mesenchymal stem cells (hMSCs) or human adult dermal fibroblasts (HDFa). For bone tissue engineering applications, we have shown that hMSCs bioprinted in TissueFab™ - Bone bioinks show an increase in osteogenic differentiation. Additionally, TissueFab™- Conductive bioinks exhibit enhanced conductivity making them attractive for bioprinting electroactive tissues such as neural or muscular tissue. TissueFab^TM bioinks provide a robust tissue-mimetic platform for microextrusion-based bioprinting various cell types with both high printability and cell viability. TissueFab^TM bioinks enable on-demand tissue printing which is a tangible step forward for addressing drug testing and tissue engineering challenges.

Advances in materials and processes are required to transform 3D printing into a manufacturing platform capable of generating complex objects with integrated function. Despite its high dimensional accuracy and resolution, excellent surface finish, versatile modification of feedstock chemistry, and low cost printers, vat polymerization 3D printing lacks the versatility to generate multimaterial objects. In this presentation, we describe a strategy to generate functional multimaterial objects by vat polymerization 3D printing that relies on controlled phase separation. Using photoresins comprising of judiciously selected components, the diffusion of phase separating components are modulated via the kinetics of gelation, the density of the polymer network and the diffusivity of materials to control the material phases spatially. The insight gained in controlling the material phases allows a rationalized approach to formulating resins to access a wide range of material morphologies for specific applications. The approach was used to fabricate dipole antenna arrays that function at 2.4 GHz, trusses that response to compression and anti-bacterial surface. Due to the universality of this approach 3D PIPS represents a powerful method to create materials with controlled sub-phases and will accelerate the adoption of vat polymerization as a viable technique to generate functional 3D objects.

The production of biomaterials for tissue and bone engineering is still a real challenge for repairing, implanting or filling damaged bone or connective tissue. Silicate-based bioceramics have excellent bioactivity and are considered promising materials for bone regeneration; however, their synthesis and design in complex geometric forms using conventional techniques is still challenging. The design of scaffolds for tissue engineering with the mechanical and microstructural properties required to promoting cell attachment, growth and new tissue formation is one of the major challenges facing researchers in the field. Their main disadvantage is their low strength and high brittleness under applied loads.
The Polymer-Derived Ceramics (PDCs) route is one of the most advantageous approaches in the manufacture of bioceramics due to the ability to control both synthesis and shaping.
In this work, we investigated 3D printing of two preceramic polymers precursors of Ca₂SiO₄ using Stereolithography (SLA). We studied the effect of the precursor’s composition and ceramic yield on the physico-chemical properties of the glass-ceramic. After pyrolysis, the scaffolds were functionalized with silver-reduced graphe oxide composites in order to assess their antibacterial activity.
The bioceramics, characterized with good mechanical strength, showed regular geometries and a high interconnected porosity (60%) with an average pore size between 200 and 400 μm. Cell viability and cytotoxicity tests showed that the scaffolds were biocampatible and nontoxic suggeting that such 3D bioceramics are suitable for tissue engineering.

Additive manufacturing is a highly advantageous manufacturing technique that enables the production of complex shapes which cannot be produced by any other technique, and it provides an opportunity for fast prototyping and personal based productions. Despite the advantages of additive ceramics manufacturing, obtaining superior mechanical properties using this technique is still a challenge because of the need to use inks with significant additive percentage. Among the other ceramic additive manufacturing techniques, extrusion-based robocasting technique offers a low-cost opportunity for the production of high-density and high strength bodies; yet, it requires strict control over the rheological behavior of the ceramic ink and the processing conditions. The increasing interest in the topic leads to significant improvements in the design of the printing parameters. However, the studies on the relation between the slurry properties, the rheological properties, the processing conditions, and the resulting properties of the green and sintered bodies are overlooked. In this study, the importance of the powder properties on the suspension rheology and the resulting properties of the extruded green and sintered bodies were investigated. Proper choice of particles, particle size and the processing additives enabled better controlled rheological properties of suspensions, smoother extrusion through narrow channels, and eliminated clogging during extrusion and crack formation in green bodies. Moreover, sintered ceramic bodies with even 200 – 300 µm size features could be produced. The sintered bodies exhibited densities above 97% of their theoretical density. In the present work, alumina was used as a model ceramic material, but the findings can certainly be extrapolated to other ceramic systems. Such findings are particularly critical for the biological, electronic and robotic applications where the mechanical requirements are demanding.

This work has been supported by Scientific Research Projects Coordination Unit of Middle East Technical University under grant number YOP-308-2018-2677.

Ultrasound is benign and can exert forces on particles and cells. A recent advance makes it possible to form high-resolution arbitrary shaped ultrasound pressure distributions [1]. By encoding the necessary phase information into the topography of a 3D-printed plate that is placed in front of a simple ultrasound transducer, it is now possible to generate sophisticated acoustic fields inside a fluid volume. We have shown that particles can be assembled into pre-defined shapes and that the particle-assemblies can be fixed to form permanent objects [2]. The advantage of directed-assembly via ultrasound fields is that the particles assemble at once to form the object. This promises a general one-shot fabrication method. Of particular interest is to apply this method to the patterning of biological cells. We could show that the acoustic amplitude distribution of a complex image, defined by the hologram, exerts forces on biological cells and the biocompatible hydrogel they are suspended in. The resultant convection flow gently delivers the suspended cells to the image plane where they form the desired pattern [3]. The hydrogel can then crosslink to immobilize the cell pattern. The use of acoustic holography for the assembly of cells structures shows great potential to grow tissue-like structures. A spatial ultrasound modulator based on microbubble arrays [4] can replace the static 3D-printed hologram and opens the possibility to form dynamic, reconfigurable assemblies.

References
[1] K. Melde, A. G. Mark, T. Qiu, P. Fischer. Nature 537, 518-522 (2016).
[2] K. Melde, E. Choi, Z. Wu, S. Palagi, T. Qiu, P. Fischer. Adv. Mater. 30:1704507 (2018).
[3] Z. Ma, A. Holle, K. Melde, T. Qiu, K. Pöppel, V. Kadiri, P. Fischer. Adv. Mater. 32:1904181 (2020).
[4] Z. Ma, K. Melde, A.G. Athanassiadis, M. Schau, H. Richter, T. Qiu, P. Fischer. Nat. Comm. 11:4537 (2020).

Ceramic additive manufacturing has the potential to be a disruptive technology enabling rapid prototyping and the production of complex geometries. A relatively new approach for ceramic part fabrication is additive manufacturing via UV-assisted direct ink writing (UV-DIW). UV photopolymerization during the print process modifies ink rheology to enable the fabrication of more complex geometries such as overhanging or spanning structures that are otherwise difficult to make with traditional DIW processes. We have developed a UV-DIW optimized ink that consists of a preceramic polymer-based system loaded with ceramic powder, containing all off the shelf components. This ink shows easy printability as well as limited shrinkage after pyrolysis, making it ideal for near net shape fabrication and rapid prototyping without the need for time consuming synthesis steps. The relationship between ink formulation, rheology, printability, as well as the structure of the final ceramic components will be presented

We have developed a laser-biomaterial interaction-based prototyping platform capable of three fabrication modes: (1) laser direct write of cells, microbeads, and other biomaterials; (2) fabrication of cell encapsulating microspheres (microcapsules); and (3) laser micromachining of substrates. Using this system, we are able to precisely place biomaterials, such as cells, into substrates with spatial constraints from laser micromachining or wholly fabricate scaffolds that are cell laden. This enables fabrication of co-cultures in almost any geometry and controlled gradients of chemical factors. In addition, the process is parallelizable, thus allowing for numerous potential bioassay applications. One such assay is a differential system for quantifying multiple outcomes in response to multiple parallel biophysicochemical cues in competition. These novel assays are complex, reproducible, and disposable microenvironments. This presentation will summarize the control integration developed for Laser Direct Write, a 2D model of laser ablation, with a computational method demonstrating preliminary results. The biofabrication methods discussed are applied to an Organ-On-a-Chip model to develop a fully automated fabrication process.