Recent advances on enhancing 3D printing quality of protein-based inks: A review.

3D printing is an additive manufacturing technology that locates constructed models with computer-controlled printing equipment. To achieve high-quality printing, the requirements on rheological properties of raw materials are extremely restrictive. Given the special structure and high modifiability under external physicochemical factors, the rheological properties of proteins can be easily adjusted to suitable properties for 3D printing. Although protein has great potential as a printing material, there are many challenges in the actual printing process. This review summarizes the technical considerations for protein-based ink 3D printing. The physicochemical factors used to enhance the printing adaptability of protein inks are discussed. The post-processing methods for improving the quality of 3D structures are described, and the application and problems of fourth dimension (4D) printing are illustrated. The prospects of 3D printing in protein manufacturing are presented to support its application in food and cultured meat. The native structure and physicochemical factors of proteins are closely related to their rheological properties, which directly link with their adaptability for 3D printing. Printing parameters include extrusion pressure, printing speed, printing temperature, nozzle diameter, filling mode, and density, which significantly affect the precision and stability of the 3D structure. Post-processing can improve the stability and quality of 3D structures. 4D design can enrich the sensory quality of the structure. 3D-printed protein products can meet consumer needs for nutritional or cultured meat alternatives.

Modulus-tunable multifunctional hydrogel ink with nanofillers for 3D-Printed soft electronics.

Seamless integration and conformal contact of soft electronics with tissue surfaces have emerged as major challenges in realizing accurate monitoring of biological signals. However, the mechanical mismatch between the electronics and biological tissues impedes the conformal interfacing between them. Attempts have been made to utilize soft hydrogels as the bioelectronic materials to realize tissue-comfortable bioelectronics. However, hydrogels have several limitations in terms of their electrical and mechanical properties. In this study, we present the development of a 3D-printable modulus-tunable hydrogel with multiple functionalities. The hydrogel has a cross-linked double network, which greatly improves its mechanical properties. Functional fillers such as XLG or functionalized carbon nanotubes (fCNT) can be incorporated into the hydrogel to provide tunable mechanics (Young's modulus of 10-300 kPa) and electrical conductivity (electrical conductivity of ∼20 S/m). The developed hydrogel exhibits stretchability (∼1000% strain), self-healing ability (within 5 min), toughness (400-731 kJ/m3) viscoelasticity, tissue conformability, and biocompatibility. Upon examining the rheological properties in the modulated region, hydrogels can be 3D printed to customize the shape and design of the bioelectronics. These hydrogels can be fabricated into ring-shaped strain sensors for wearable sensor applications.

Graphene derivative-based ink advances inkjet printing technology for fabrication of electrochemical sensors and biosensors.

The field of biosensing would significantly benefit from a disruptive technology enabling flexible manufacturing of uniform electrodes. Inkjet printing holds promise for this, although realizing full electrode manufacturing with this technology remains challenging. We introduce a nitrogen-doped carboxylated graphene ink (NGA-ink) compatible with commercially available printing technologies. The water-based and additive-free NGA-ink was utilized to produce fully inkjet-printed electrodes (IPEs), which demonstrated successful electrochemical detection of the important neurotransmitter dopamine. The cost-effectiveness of NGA-ink combined with a total cost per electrode of $0.10 renders it a practical solution for customized electrode manufacturing. Furthermore, the high carboxyl group content of NGA-ink (13 wt%) presents opportunities for biomolecule immobilization, paving the way for the development of advanced state-of-the-art biosensors. This study highlights the potential of NGA inkjet-printed electrodes in revolutionizing sensor technology, offering an affordable, scalable alternative to conventional electrochemical systems.

Recent Developments in Bio-Ink Formulations Using Marine-Derived Biomaterials for Three-Dimensional (3D) Bioprinting.

3D bioprinting is a disruptive, computer-aided, and additive manufacturing technology that allows the obtention, layer-by-layer, of 3D complex structures. This technology is believed to offer tremendous opportunities in several fields including biomedical, pharmaceutical, and food industries. Several bioprinting processes and bio-ink materials have emerged recently. However, there is still a pressing need to develop low-cost sustainable bio-ink materials with superior qualities (excellent mechanical, viscoelastic and thermal properties, biocompatibility, and biodegradability). Marine-derived biomaterials, including polysaccharides and proteins, represent a viable and renewable source for bio-ink formulations. Therefore, the focus of this review centers around the use of marine-derived biomaterials in the formulations of bio-ink. It starts with a general overview of 3D bioprinting processes followed by a description of the most commonly used marine-derived biomaterials for 3D bioprinting, with a special attention paid to chitosan, glycosaminoglycans, alginate, carrageenan, collagen, and gelatin. The challenges facing the application of marine-derived biomaterials in 3D bioprinting within the biomedical and pharmaceutical fields along with future directions are also discussed.

From voice to ink (Vink): development and assessment of an automated, free-of-charge transcription tool.

BACKGROUND: Verbatim transcription of qualitative audio data is a cornerstone of analytic quality and rigor, yet the time and energy required for such transcription can drain resources, delay analysis, and hinder the timely dissemination of qualitative insights. In recent years, software programs have presented a promising mechanism to accelerate transcription, but the broad application of such programs has been constrained due to expensive licensing or "per-minute" fees, data protection concerns, and limited availability of such programs in many languages. In this article, we outline our process of adapting a free, open-source, speech-to-text algorithm (Whisper by OpenAI) into a usable and accessible tool for qualitative transcription. Our program, which we have dubbed "Vink" for voice to ink, is available under a permissive open-source license (and thus free of cost). RESULTS: We conducted a proof-of-principle assessment of Vink's performance in transcribing authentic interview audio data in 14 languages. A majority of pilot-testers evaluated the software performance positively and indicated that they were likely to use the tool in their future research. Our usability assessment indicates that Vink is easy-to-use, and we performed further refinements based on pilot-tester feedback to increase user-friendliness. CONCLUSION: With Vink, we hope to contribute to facilitating rigorous qualitative research processes globally by reducing time and costs associated with transcription and by expanding free-of-cost transcription software availability to more languages. With Vink running on standalone computers, data privacy issues arising within many other solutions do not apply.

Gelatin Methacryloyl (GelMA)-Based Biomaterial Inks: Process Science for 3D/4D Printing and Current Status.

Tissue engineering for injured tissue replacement and regeneration has been a subject of investigation over the last 30 years, and there has been considerable interest in using additive manufacturing to achieve these goals. Despite such efforts, many key questions remain unanswered, particularly in the area of biomaterial selection for these applications as well as quantitative understanding of the process science. The strategic utilization of biological macromolecules provides a versatile approach to meet diverse requirements in 3D printing, such as printability, buildability, and biocompatibility. These molecules play a pivotal role in both physical and chemical cross-linking processes throughout the biofabrication, contributing significantly to the overall success of the 3D printing process. Among the several bioprintable materials, gelatin methacryloyl (GelMA) has been widely utilized for diverse tissue engineering applications, with some degree of success. In this context, this review will discuss the key bioengineering approaches to identify the gelation and cross-linking strategies that are appropriate to control the rheology, printability, and buildability of biomaterial inks. This review will focus on the GelMA as the structural (scaffold) biomaterial for different tissues and as a potential carrier vehicle for the transport of living cells as well as their maintenance and viability in the physiological system. Recognizing the importance of printability toward shape fidelity and biophysical properties, a major focus in this review has been to discuss the qualitative and quantitative impact of the key factors, including microrheological, viscoelastic, gelation, shear thinning properties of biomaterial inks, and printing parameters, in particular, reference to 3D extrusion printing of GelMA-based biomaterial inks. Specifically, we emphasize the different possibilities to regulate mechanical, swelling, biodegradation, and cellular functionalities of GelMA-based bio(material) inks, by hybridization techniques, including different synthetic and natural biopolymers, inorganic nanofillers, and microcarriers. At the close, the potential possibility of the integration of experimental data sets and artificial intelligence/machine learning approaches is emphasized to predict the printability, shape fidelity, or biophysical properties of GelMA bio(material) inks for clinically relevant tissues.

Smart nano-inks based on natural food colorant for screen-printing of dynamic shelf life of shrimp.

Intelligent packaging embedded with food freshness indicators can monitor food quality and be deployed for food safety and cutting food waste. The innovative nano-inks for dynamic shelf-life printing based on natural food colorant with application in real-time monitoring of shrimp freshness were prepared. Co-assembly of saffron petal anthocyanin (SPA) with hydrophobic curcumin (Cur) into chitin nano-scaffold (particle sizes around 26 ± 8 nm) could deliver hindering SPA leaching, confirmed by FT-IR, FE-SEM, AFM, and color stability test. The best response to pH-sensitivity was found in a ratio of (1:4) Cur/SPA (30% (v/w) in ChNFs that was correlated with the chemical and microbial changes of shrimp during shrimp freshness. However, smart screen-printed inks signified higher responsiveness to pH changes than FFI films. Therefore, smart-printed indicators introduced the excellent potential for a short response time, easy, cost-effective, eco-friendly, co-assembly, great color stabilities, and lifetime for nondestructively freshness monitoring foods and supplements.

Rheological insights into 3D printing of drug products: Drug nanocrystal-poloxamer gels for semisolid extrusion.

The importance of ink rheology to the outcome of 3D printing is well recognized. However, rheological properties of printing inks containing drug nanocrystals have not been widely investigated. Therefore, the objective of this study was to establish a correlation between the composition of nanocrystal printing ink, the ink rheology, and the entire printing process. Indomethacin was used as a model poorly soluble drug to produce nanosuspensions with improved solubility properties through particle size reduction. The nanosuspensions were further developed into semisolid extrusion 3D printing inks with varying nanocrystal and poloxamer 407 concentrations. Nanocrystals were found to affect the rheological properties of the printing inks both by being less self-supporting and having higher yielding resistances. During printing, nozzle blockages occurred. Nevertheless, all inks were found to be printable. Finally, the rheological properties of the inks were successfully correlated with various printing and product properties. Overall, these experiments shed new light on the rheological properties of printing inks containing nanocrystals.

Time-temperature indicator of hydroxyethyl cellulose ink labels for assessing pork freshness.

Pork is widely consumed worldwide, and many consumers now utilize sensory evaluation techniques to determine the freshness of pork when buying it. A color-changing ink label utilizing bromocresol purple (BCP) and N-hydroxyphthalimide (NHPI) had been created to help consumers better and more rapidly determine the freshness of pork while it is stored. The ink was easy to prepare and could be readily transferred to A4 paper using screen printing technology. This study delved deeper into the impact of hydroxyethyl cellulose (HEC) on the functional properties of inks to enhance printing performance. The experiment demonstrated that a 1 % mass fraction of HEC improved thixotropy and facilitated the even distribution of ink on A4 paper, as confirmed by scanning electron microscopy. Screen-printed labels with varying concentrations displayed distinct color change rates when stored at different temperatures, indicating their capability to assess pork freshness. FT-IR, laboratory, and stability tests verified the ink's exceptional color change capabilities and printing attributes. An analysis using the Arrhenius equation revealed a substantial synergistic effect between BCP and NHPI, resulting in improved sensitivity and accuracy of the ink. This study offers a practical and feasible method to monitor the storage quality of pork effectively.

Effect of a collagen peptide-fish oil high internal phase emulsion on the printability and gelation of 3D-printed surimi gel inks.

The effect of a high internal phase emulsion (HIPE) on three-dimensional-printed surimi gel inks was studied. Increasing the concentration of collagen peptide decreased the particle size of HIPE droplets and improved the viscoelasticity and stability. For example, when the collagen peptide concentration was 5 wt%, the viscoelasticity of the HIPE was high, as indicated by the presence of small and uniform particles, which formed a monolayer in the outer layer of the oil droplets to form stable a HIPE. A HIPE was used as the filling material to fill the surimi gel network, which reduced the porosity of the network. Surimi protein and peptides have dual emulsifying effects on the stabilization of oil. After adding the emulsion, the texture, gel properties and rheological properties of the surimi were reduced, and its printing adaptability was improved. This study provides new ideas for the production of surimi and its application in 3D printing.

Toughening 3D printed biomimetic hydroxyapatite scaffolds: Polycaprolactone-based self-hardening inks.

The application of 3D printing to calcium phosphates has opened unprecedented possibilities for the fabrication of personalized bone grafts. However, their biocompatibility and bioactivity are counterbalanced by their high brittleness. In this work we aim at overcoming this problem by developing a self-hardening ink containing reactive ceramic particles in a polycaprolactone solution instead of the traditional approach that use hydrogels as binders. The presence of polycaprolactone preserved the printability of the ink and was compatible with the hydrolysis-based hardening process, despite the absence of water in the ink and its hydrophobicity. The microstructure evolved from a continuous polymeric phase with loose ceramic particles to a continuous network of hydroxyapatite nanocrystals intertwined with the polymer, in a configuration radically different from the polymer/ceramic composites obtained by fused deposition modelling. This resulted in the evolution from a ductile behavior, dominated by the polymer, to a stiffer behavior as the ceramic phase reacted. The polycaprolactone binder provides two highly relevant benefits compared to hydrogel-based inks. First, the handleability and elasticity of the as-printed scaffolds, together with the proven possibility of eliminating the solvent, opens the door to implanting the scaffolds freshly printed once lyophilized, while in a ductile state, and the hardening process to take place inside the body, as in the case of calcium phosphate cements. Second, even with a hydroxyapatite content of more than 92 wt.%, the flexural strength and toughness of the scaffolds after hardening are twice and five times those of the all-ceramic scaffolds obtained with the hydrogel-based inks, respectively. STATEMENT OF SIGNIFICANCE: Overcoming the brittleness of ceramic scaffolds would extend the applicability of synthetic bone grafts to high load-bearing situations. In this work we developed a 3D printing ink by replacing the conventional hydrogel binder with a water-free polycaprolactone solution. The presence of polycaprolactone not only enhanced significantly the strength and toughness of the scaffolds while keeping the proportion of bioactive ceramic phase larger than 90 wt.%, but it also conferred flexibility and manipulability to the as-printed scaffolds. Since they are able to harden upon contact with water under physiological conditions, this opens up the possibility of implanting them immediately after printing, while they are still in a ductile state, with clear advantages for fixation and press-fit in the bone defect.

Aloe vera-based biomaterial ink for 3D bioprinting of wound dressing constructs.

This study emphasizes the development of a multifunctional biomaterial ink for wound healing constructs. The biomaterial ink benefits from Aloe vera's intrinsic biocompatible, biodegradable, antioxidant, antimicrobial, anti-inflammatory, and immunomodulatory attributes, thus alleviating the need for supplementary substances employed to combat infections and stimulate tissue regeneration. Moreover, this biomaterial ink seeks to address the scarcity of standardized printable materials possessing adequate biocompatibility and physicochemical properties, which hinder its widespread clinical adoption. The biomaterial ink was synthesized via ionic crosslinking to enhance its rheological and mechanical characteristics. The findings revealed that Aloe vera substantially boosted the hydrogel's viscoelastic behavior, enabling superior compressive modulus and the extrusion of fine filaments. The bioprinted constructs exhibited desirable resolution and mechanical strength while displaying a porous microstructure analogous to the native extracellular matrix. Biological response demonstrated no detrimental impact on stem cell viability upon exposure to the biomaterial ink, as confirmed by live/dead assays. These outcomes validate the potential of the developed biomaterial ink as a resource for the bioprinting of wound dressings that effectively foster cellular proliferation, thereby promoting enhanced wound healing by leveraging Aloe vera's inherent properties.

Innovations in hydrogel-based manufacturing: A comprehensive review of direct ink writing technique for biomedical applications.

Direct ink writing (DIW) stands as a pioneering additive manufacturing technique that holds transformative potential in the field of hydrogel fabrication. This innovative approach allows for the precise deposition of hydrogel inks layer by layer, creating complex three-dimensional structures with tailored shapes, sizes, and functionalities. By harnessing the versatility of hydrogels, DIW opens up possibilities for applications spanning from tissue engineering to soft robotics and wearable devices. This comprehensive review investigates DIW as applied to hydrogels and its multifaceted applications. The paper introduces a diverse range of printing techniques while providing a thorough exploration of DIW for hydrogel-based printing. The investigation aims to explain the progress made, challenges faced, and potential trajectories that lie ahead for DIW in hydrogel-based manufacturing. The fundamental principles underlying DIW are carefully examined, specifically focusing on rheological attributes and printing parameters, prompting a comprehensive survey of the wide variety of hydrogel materials. These encompass both natural and synthetic variations, all of which can be effectively harnessed for this purpose. Furthermore, the review explores the latest applications of DIW for hydrogels in biomedical areas, with a primary focus on tissue engineering, wound dressing, and drug delivery systems. The document not only consolidates the existing state of DIW within the context of hydrogel-based manufacturing but also charts potential avenues for further research and innovative breakthroughs.

Tattoo Pigment Biokinetics in vivo in a 28-Day Porcine Model: Elements Undergo Fast Distribution to Lymph Nodes and Reach Steady State after 7 Days.

INTRODUCTION: Pigments of tattoo inks may over time migrate to other parts of the body. Inks kinetics are still poorly understood and little studied. The aim of this first study was to investigate the kinetics of tattoo inks pigment in tattooed porcine skin, which is closer to human skin than mouse skin studied in the past. METHODS: Three animals were tattooed on the inner thigh and one animal served as untreated control. Skin biopsies were taken on days 7, 14, and 28 after tattooing. Animals were sacrificed on day 28 and homogenate samples of the liver, spleen, kidney, and brain, as well the local lymph nodes were prepared. All samples were analyzed for ink components using inductively coupled plasma-mass spectrometry. The ink itself was characterized by dynamic light scattering and matrix-assisted laser desorption-ionization mass analysis. RESULTS: Titanium (212 g/kg), copper (6 mg/kg), aluminum (1 mg/kg), zirconium (1 mg/kg), and chromium (3 mg/kg) were found in the ink. Significant deposits of ink elements were detected in the tattooed skin when compared to non-tattooed skin from the same animal (mean ± standard deviation: titanium 240 ± 81 mg/kg, copper 95 ± 39 mg/kg, aluminum 115 ± 63 mg/kg, zirconium 23 ± 12 mg/kg, and chromium 1.0 ± 0.2 mg/kg; p < 0.05). Lymph node concentrations of titanium, copper, aluminum, zirconium, and chromium were 42 ± 2 mg/kg, 69 ± 25 mg/kg, 49 ± 18 mg/kg, 0.3 ± 0.2 mg/kg, 0.5 ± 0.2 mg/kg, respectively. CONCLUSION: Deposits in skin were unchanged from days 7-28 indicating no redistribution or elimination. No significant deposits of ink elements were found in the liver, spleen, kidney, and brain. In conclusion, our findings confirmed distribution of elements from tattoos to regional lymph nodes, but neither to excretory organs, e.g., liver and kidney, nor to spleen and brain. Thus systemic internal organ exposure was not found.

What's in My Ink: An Analysis of Commercial Tattoo Ink on the US Market.

As tattoos continue to rise in popularity, the demand for tattoo ink has surged. Historically, tattoo inks have been underregulated in the US market. This study analyzes inks from nine different brands that are common in the United States, ranging from major to small manufacturers. Out of 54 inks, 45 contained unlisted additives and/or pigments. Major, unlisted adulterants include poly(ethylene glycol), propylene glycol, and higher alkanes. Many of the adulterants pose possible allergic or other health risks. Taken together, the results from this study highlight the potential for a significant issue around inaccurate tattoo ink labeling in the United States.

Effect of polymethyl methacrylate on in situ patterning of perovskite quantum dots by inkjet printing.

The preparation of perovskite quantum dots (PQDs) using an in situ inkjet printing method is beneficial for improving the problems of aggregation and photoluminescence (PL) quenching during long-term storage. However, the stability of PQDs prepared using this method is still not ideal, and the morphology of in situ-printed patterns needs to be optimized. To address these problems, this study introduced polymethyl methacrylate (PMMA) into the process of in situ inkjet printing of PQDs and explored the effect of PMMA on the in situ patterning effect of PQDs. The results showed that using a mixed precursor solution containing a small amount of PMMA as the printing ink can slow down the shrinkage process of ink droplets and improve the uniformity of film formation. As the printing substrate, PMMA provided a suitable high-viscosity environment for the in situ growth of PQDs. This could effectively suppress the coffee ring effect. In addition, the interaction between the C=O=C group in PMMA and metal ion Pb2+ in the CsPbBr3 precursor molecules was favourable to enhancing the density of PQDs. The prepared PMMA-coated CsPbBr3 quantum dots (QDs) pattern had high stability and could maintain at 90.08% PL intensity after 1 week of exposure to air.

Novel-Ink-Based Direct Ink Writing of Ti6Al4V Scaffolds with Sub-300 µm Structural Pores for Superior Cell Proliferation and Differentiation.

Ti6Al4V scaffolds with pore sizes between 300 and 600 µm are deemed suitable for bone tissue engineering. However, a significant proportion of human bone pores are smaller than 300 µm, playing a crucial role in cell proliferation, differentiation, and bone regeneration. Ti6Al4V scaffolds with these small-sized pores are not successfully fabricated, and their cytocompatibility remains unknown. The study presents a novel ink formula specifically tailored for fabricating Ti6Al4V scaffolds featuring precise and unobstructed sub-300 µm structural pores, achieved by investigating the rheological properties and printability of five inks containing 60-77.5 vol% Ti6Al4V powders and bisolvent binders. Ti6Al4V scaffolds with 50-600 µm pores are fabricated via direct ink writing and subjected to in vitro assays with MC3T3-E1 and bone marrow mesenchymal stem cells. The 100 µm pore-sized scaffolds exhibit the highest cell adhesion and proliferation capacity based on live/dead assay, FITC-phalloidin/4',6-diamidino-2-phenylindole staining, and cell count kit 8 assay. The alizarin red staining, real-time quantitative PCR assay, and immunocytochemical staining demonstrate the superior osteogenic differentiation potential of 100 and 200 µm pore-sized scaffolds. The importance of sub-300 µm structrual pores is highlighted, redefining the optimal pore size for Ti6Al4V scaffolds and advancing bone tissue engineering and clinical medicine development.

From biomaterials to biotherapy: cuttlefish ink with protoporphyrin IX nanoconjugates for synergistic sonodynamic-photothermal therapy.

Biologically produced nanomaterials capable of therapeutic purposes have received increasing interest in tumor therapy because of their intrinsic biocompatibility. In this study, we made cuttlefish ink (extracted from cuttlefish) and protoporphyrin IX (PpIX) nanoconjugates (CIPs) where PpIX was an endogenous organic compound. In the case of CIPs, PpIX could be triggered by ultrasound (US) for sonodynamic therapy (SDT), and the cuttlefish ink could be excited by a near-infrared laser for photothermal therapy (PTT). Thereafter, tumor growth was greatly inhibited through synergistic SDT-PTT in comparison to single SDT or PTT. In addition, in vivo administration of CIPs showed no noticeable side effects for mouse blood and chief organs, providing an effective strategy for developing biologically produced biomaterials and using them for biotherapy.

Three-Dimensional Extrusion Printed Urinary Specific Grafts: Mechanistic Insights into Buildability and Biophysical Properties.

The compositional formulations and the optimization of process parameters to fabricate hydrogel scaffolds with urological tissue-mimicking biophysical properties are not yet extensively explored, including a comprehensive assessment of a spectrum of properties, such as mechanical strength, viscoelasticity, antimicrobial property, and cytocompatibility. While addressing this aspect, the present work provides mechanistic insights into process science, to produce shape-fidelity compliant alginate-based biomaterial ink blended with gelatin and synthetic nanocellulose. The composition-dependent pseudoplasticity, viscoelasticity, thixotropy, and gel stability over a longer duration in physiological context have been rationalized in terms of intermolecular hydrogen bonding interactions among the biomaterial ink constituents. By varying the hybrid hydrogel ink composition within a narrow compositional window, the resulting hydrogel closely mimics the natural urological tissue-like properties, including tensile stretchability, compressive strength, and biophysical properties. Based on the printability assessment using a critical analysis of gel strength, we have established the buildability of the acellular hydrogel ink and have been successful in fabricating shape-fidelity compliant urological patches or hollow cylindrical grafts using 3D extrusion printing. Importantly, the new hydrogel formulations with good hydrophilicity, support fibroblast cell proliferation and inhibit the growth of Gram-negative E. coli bacteria. These attributes were rationalized in terms of nanocellulose-induced physicochemical changes on the scaffold surface. Taken together, the present study uncovers the process-science-based understanding of the 3D extrudability of the newly formulated alginate-gelatin-nanocellulose-based hydrogels with urological tissue-specific biophysical, cytocompatibility, and antibacterial properties.