Bioink properties before, during and after 3D bioprinting

Bioprinting is a process based on additive manufacturing from materials containing living cells. These materials, often referred to as bioink, are based on cytocompatible hydrogel precursor formulations, which gel in a manner compatible with different bioprinting approaches. The bioink properties before, during and after gelation are essential for its printability, comprising such features as achievable structural resolution, shape fidelity and cell survival. However, it is the final properties of the matured bioprinted tissue construct that are crucial for the end application. During tissue formation these properties are influenced by the amount of cells present in the construct, their proliferation, migration and interaction with the material. A calibrated computational framework is able to predict the tissue development and maturation and to optimize the bioprinting input parameters such as the starting material, the initial cell loading and the construct geometry. In this contribution relevant bioink properties are reviewed and discussed on the example of most popular bioprinting approaches. The effect of cells on hydrogel processing and vice versa is highlighted. Furthermore, numerical approaches were reviewed and implemented for depicting the cellular mechanics within the hydrogel as well as for prediction of mechanical properties to achieve the desired hydrogel construct considering cell density, distribution and material-cell interaction

Flow-based fabrication: An integrated computational workflow for design and digital additive manufacturing of multifunctional heterogeneously structured objects

Structural hierarchy and material organization in design are traditionally achieved by combining discrete homogeneous parts into functional assemblies where the shape or surface is the determining factor in achieving function. In contrast, biological structures express higher levels of functionality on a finer scale through volumetric cellular constructs that are heterogeneous and complex. Despite recent advancements in additive manufacturing of functionally graded materials, the limitations associated with computational design and digital fabrication of heterogeneous materials and structures frame and limit further progress. Conventional computer-aided design tools typically contain geometric and topologic data of virtual constructs, but lack robust means to integrate material composition properties within virtual models. We present a seamless computational workflow for the design and direct digital fabrication of multi-material and multi-scale structured objects. The workflow encodes for and integrates domain-specific meta-data relating to local, regional and global feature resolution of heterogeneous material organizations. We focus on water-based materials and demonstrate our approach by additively manufacturing diverse constructs associating shape-informing variable flow rates and material properties to mesh-free geometric primitives. The proposed workflow enables virtual-to-physical control of constructs where structural, mechanical and optical gradients are achieved through a seamless design-to-fabrication tool with localized control. An enabling technology combining a robotic arm and a multi-syringe multi nozzle deposition system is presented. Proposed methodology is implemented and full-scale demonstrations are included

Printed and drawn flexible electronics based on cellulose nanocomposites

Sustainability, flexibility, and low-power consumption are key features to meet the growing re- quirements of simplicity and multifunctionality of low-cost, disposable/recyclable smart electronic -of- -based composites hold po- tential to fulfill such demands when explored as substrate and/or electrolyte-gate, or as active channel layer on printed transistors and integrated circuits based on ionic responses (iontronics). In this work, a new generation of reusable, healable and recyclable regenerated cellulose hydro- gels with high ionic conductivity and conformability, capable of being provided in the form of stick- ers, are demonstrated. These hydrogels are obtained from a simple, fast, low-cost, and environ- mental-friendly aqueous alkali salt/urea dissolution method of native cellulose, combined with eration and simultaneous ion incorporation with acetic acid. Their electrochemical properties can be also merged with the mechanical robustness, thermal resistance, transparency, and smooth- - strate. Beyond gate dielectrics, a water-based screen-printable ink, composed of CMC binder and com- mercial zinc oxide (ZnO) semiconducting nanoparticles, was formulated. The ink enables the printing of relatively smooth and densely packed films on office paper with semiconducting func- tionality at room temperature. The rather use of porous ZnO nanoplates is beneficial to form per- colative pathways at lower contents of functional material, at the cost of rougher surfaces. The engineered cellulose composites are successfully integrated into flexible, recyclable, low- voltage (<3.5 V), printed electrolyte-gated office paper or on the ionically modified nanopaper. Ubiquitous calligraphy accessories are used -the- out on the target substrate, where are already printed the devices. Such concept paves the way for a worldwide boom of creativity, where we can freely create personal electronic kits, while having fun at it and without generating waste.Sustentabilidade, flexibilidade e baixo consumo energético são características chave para atender aos crescentes requisitos de simplicidade e multifuncionalidade de sistemas eletrónicos inteligentes de baixo custo, das- Compósitos à base de celulose têm potencial para atender a tais necessidades quando explora- dos como substrato e/ou porta-de-eletrólito ou como camada de canal ativo em transístores impressos e circuitos integrados baseados em respostas iónicas (iontronics). Neste trabalho, é demonstrada uma nova geração de hidrogéis reutilizáveis, reparáveis e recicláveis baseados em celulose regenerada, que apresentam alta condução iónica e conformabilidade, podendo ser fornecidos na forma de adesivos. Estes hidrogéis são obtidos a partir de um método simples, rápido, barato e amigo do ambiente que permite a dissolução de celulose nativa em soluções aquosas com mistura de sal alcalino e ureia, combinado com carboximetil celulose (CMC) para melhorar a sua robustez, seguido da regeneração e simultâneo enriquecimento iónico com ácido acético. As suas propriedades eletroquímicas podem ser combinadas com a inbase de celulose micro/nanofibrilada para obter um substrato eletrolítico semelhante a papel. Para além de portas-dielétricas, foi formulada uma tinta aquosa compatível com serigrafia, composta por CMC como espessante e nanopartículas semicondutoras de ZnO. A tinta permite a impressão de filmes pouco rugosos e densamente percolados sobre papel de escritório, e com funcionalidade semicondutora à temperatura ambiente. O uso alternativo de nanoplacas porosas de ZnO é benéfico para criar caminhos percolativos com menores teores de material funcional, apesar de se obter filmes rugosos. Os compósitos à base celulose foram integrados com sucesso em transístores e portas lógicas porta-eletrolítica, os quais foram impressos em papel de escritório ou no "nanopapel" iconicamente modificado. Acessórios de caligrafia permitem a fácil e rápida padronização de pistas condutoras/resistivas, desenhando-as no substrato alvo, onde estão impressos os dispositivos. Este conceito despoleta um mundo criativo, onde é possível criar livremente kits eletrónicos customizados de forma divertida e sem gerar resíduos

SWELLING INDUCED DEFORMATION OF THERMALLY RESPONSIVE HYDROGELS

Hydrogels are crosslinked polymeric networks imbibed with aqueous solutions. They undertake dramatic volume changes through swelling and deswelling processes, which can be stimulated by factors like temperature, pH or different chemicals. These unique properties render hydrogels particularly interesting for shape morphing related applications. In this thesis, we focus on the swelling induced deformation of thermally responsive hydrogels with lower critical solution temperatures (LCSTs), including poly(N-isopropylacrylamide) (PNIPAm) and poly(N,N-diethylacrylamide) (PDEAm). Particularly, benzophenone containing monomers are copolymerized with NIPAm or DEAm to create photocrosslinkable temperature-responsive polymers, which allows fabrication of hydrogels with controlled shapes and crosslinking densities by photolithography. Based on this material system, non-uniform swelling is patterned to cause complicated 3D deformation of hydrogel sheets in Chapter 2. Relying on differential geometry, a collection of 3D shapes is successfully programmed by prescribing Gaussian curvature with in-plane swelling metrics. However, this methodology does not have control over the buckling direction of each local maximum in the swelling metric and cannot distinguish different isometric shapes. Thus, modest swelling gradients through the gel sheet thickness are introduced via absorption of light to provide preferential buckling directions for each local maximum. In the example of double sinusoidal surfaces, the buckling direction of each repeating unit is effectively controlled with double-sided photolithography. Next, we emphasize the actuation behaviors of these thermally responsive hydrogels. In addition to thermal actuation, photo actuation of a PNIPAm based hydrogel bilayer structure is demonstrated by the incorporation of photothermal effects from plasmonic nanoparticles. Especially, waveguided light is used here, providing remotely controllable actuation that does not require line-of-sight access

Bioink properties before, during and after 3D bioprinting

Bioprinting is a process based on additive manufacturing from materials containing living cells. These materials, often referred to as bioink, are based on cytocompatible hydrogel precursor formulations, which gel in a manner compatible with different bioprinting approaches. The bioink properties before, during and after gelation are essential for its printability, comprising such features as achievable structural resolution, shape fidelity and cell survival. However, it is the final properties of the matured bioprinted tissue construct that are crucial for the end application. During tissue formation these properties are influenced by the amount of cells present in the construct, their proliferation, migration and interaction with the material. A calibrated computational framework is able to predict the tissue development and maturation and to optimize the bioprinting input parameters such as the starting material, the initial cell loading and the construct geometry. In this contribution relevant bioink properties are reviewed and discussed on the example of most popular bioprinting approaches. The effect of cells on hydrogel processing and vice versa is highlighted. Furthermore, numerical approaches were reviewed and implemented for depicting the cellular mechanics within the hydrogel as well as for prediction of mechanical properties to achieve the desired hydrogel construct considering cell density, distribution and material–cell interaction

Bioanalytical applications of microfluidic devices

The first part of the thesis describes a new patterning technique--microfluidic contact printing--that combines several of the desirable aspects of microcontact printing and microfluidic patterning and addresses some of their important limitations through the integration of a track-etched polycarbonate (PCTE) membrane. Using this technique, biomolecules (e.g., peptides, polysaccharides, and proteins) were printed in high fidelity on a receptor modified polyacrylamide hydrogel substrate. The patterns obtained can be controlled through modifications of channel design and secondary programming via selective membrane wetting. The protocols support the printing of multiple reagents without registration steps and fast recycle times. The second part describes a non-enzymatic, isothermal method to discriminate single nucleotide polymorphisms (SNPs). SNP discrimination using alkaline dehybridization has long been neglected because the pH range in which thermodynamic discrimination can be done is quite narrow. We found, however, that SNPs can be discriminated by the kinetic differences exhibited in the dehybridization of PM and MM DNA duplexes in an alkaline solution using fluorescence microscopy. We combined this method with multifunctional encoded hydrogel particle array (fabricated by stop-flow lithography) to achieve fast kinetics and high versatility. This approach may serve as an effective alternative to temperature-based method for analyzing unamplified genomic DNA in point-of-care diagnostic

3D Cell Printed Tissue Analogues: A New Platform for Theranostics

Stem cell theranostics has received much attention for noninvasively monitoring and tracing transplanted therapeutic stem cells through imaging agents and imaging modalities. Despite the excellent regenerative capability of stem cells, their efficacy has been limited due to low cellular retention, low survival rate, and low engraftment after implantation. Three-dimensional (3D) cell printing provides stem cells with the similar architecture and microenvironment of the native tissue and facilitates the generation of a 3D tissue-like construct that exhibits remarkable regenerative capacity and functionality as well as enhanced cell viability. Thus, 3D cell printing can overcome the current concerns of stem cell therapy by delivering the 3D construct to the damaged site. Despite the advantages of 3D cell printing, the in vivo and in vitro tracking and monitoring of the performance of 3D cell printed tissue in a noninvasive and real-time manner have not been thoroughly studied. In this review, we explore the recent progress in 3D cell technology and its applications. Finally, we investigate their potential limitations and suggest future perspectives on 3D cell printing and stem cell theranostics.116Nsciescopu

Multizone Paper Platform for 3D Cell Cultures

In vitro 3D culture is an important model for tissues in vivo. Cells in different locations of 3D tissues are physiologically different, because they are exposed to different concentrations of oxygen, nutrients, and signaling molecules, and to other environmental factors (temperature, mechanical stress, etc). The majority of high-throughput assays based on 3D cultures, however, can only detect the average behavior of cells in the whole 3D construct. Isolation of cells from specific regions of 3D cultures is possible, but relies on low-throughput techniques such as tissue sectioning and micromanipulation. Based on a procedure reported previously (“cells-in-gels-in-paper” or CiGiP), this paper describes a simple method for culture of arrays of thin planar sections of tissues, either alone or stacked to create more complex 3D tissue structures. This procedure starts with sheets of paper patterned with hydrophobic regions that form 96 hydrophilic zones. Serial spotting of cells suspended in extracellular matrix (ECM) gel onto the patterned paper creates an array of 200 micron-thick slabs of ECM gel (supported mechanically by cellulose fibers) containing cells. Stacking the sheets with zones aligned on top of one another assembles 96 3D multilayer constructs. De-stacking the layers of the 3D culture, by peeling apart the sheets of paper, “sections” all 96 cultures at once. It is, thus, simple to isolate 200-micron-thick cell-containing slabs from each 3D culture in the 96-zone array. Because the 3D cultures are assembled from multiple layers, the number of cells plated initially in each layer determines the spatial distribution of cells in the stacked 3D cultures. This capability made it possible to compare the growth of 3D tumor models of different spatial composition, and to examine the migration of cells in these structures

Classification of analytics, sensorics, and bioanalytics with polyelectrolyte multilayer capsules

Polyelectrolyte multilayer (PEM) capsules, constructed by LbL (layer-by-layer)-adsorbing polymers on sacrificial templates, have become important carriers due to multifunctionality of materials adsorbed on their surface or encapsulated into their interior. They have been also been used broadly used as analytical tools. Chronologically and traditionally, chemical analytics has been developed first, which has long been synonymous with all analytics. But it is not the only development. To the best of our knowledge, a summary of all advances including their classification is not available to date. Here, we classify analytics, sensorics, and biosensorics functionalities implemented with polyelectrolyte multilayer capsules and coated particles according to the respective stimuli and application areas. In this classification, three distinct categories are identified: (I) chemical analytics (pH; K+, Na+, and Pb2+ ion; oxygen; and hydrogen peroxide sensors and chemical sensing with surface-enhanced Raman scattering (SERS)); (II) physical sensorics (temperature, mechanical properties and forces, and osmotic pressure); and (III) biosensorics and bioanalytics (fluorescence, glucose, urea, and protease biosensing and theranostics). In addition to this classification, we discuss also principles of detection using the above-mentioned stimuli. These application areas are expected to grow further, but the classification provided here should help (a) to realize the wealth of already available analytical and bioanalytical tools developed with capsules using inputs of chemical, physical, and biological stimuli and (b) to position future developments in their respective fields according to employed stimuli and application areas