Targeted micro-heterogeneity in bioinks allows for 3D printing of complex constructs with improved resolution and cell viability
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Three-dimensional bioprinting is an evolving versatile technique for biomedical applications. Ideal bioinks have complex micro-environment that mimic human tissue, allow for good printing quality and provide high cell viability after printing. Here we present two strategies for enhancing gelatin-based bioinks heterogeneity on a 1-100µm length scale resulting in superior printing quality and high cell viability. A thorough spatial and micro-mechanical characterization of swollen hydrogel heterogeneity was done using multiple particle tracking microrheology. When poly(vinyl alcohol) is added to homogeneous gelatin gels, viscous inclusions are formed due to micro-phase separation. This phenomenon leads to pronounced slip and superior printing quality of complex 3D constructs as well as high human hepatocellular carcinoma (HepG2) and normal human dermal fibroblast (NHDF) cell viability due to reduced shear damage during extrusion. Similar printability and cell viability results are obtained with gelatin/nanoclay composites. The formation of polymer/nanoclay clusters reduces the critical stress of gel fracture, which facilitates extrusion, thus enhancing printing quality and cell viability. Targeted introduction of micro-heterogeneities in bioinks through micro-phase separation is an effective technique for high resolution 3D printing of complex constructs with high cell viability. The size of the heterogeneities, however, has to be substantially smaller than the desired feature size in order to achieve good printing quality.Keywords:
Gelatin
Viability assay
Detecting cell viability is crucial in research involving the precancerous discovery of abnormal cells, the evaluation of treatments, and drug toxicity testing. Although conventional methods afford cumulative results regarding cell viability based on a great number of cells, they do not permit investigating cell viability at the single-cell level. In response, we rationally designed and synthesized a fluorescent probe, PCV-1, to visualize cell viability under the super-resolution technology of structured illumination microscopy. Given its sensitivity to mitochondrial membrane potential and affinity to DNA, PCV-1's ability to stain mitochondria and nucleoli was observed in live and dead cells, respectively. During cell injury induced by drug treatment, PCV-1's migration from mitochondria to the nucleolus was dynamically visualized at the single-cell level. By extension, harnessing PCV-1's excellent photostability and signal-to-noise ratio and by comparing the fluorescence intensity of the two organelles, mitochondria and nucleoli, we developed a powerful analytical assay named organelle ratiometric probing (ORP) that we applied to quantitatively analyze and efficiently assess the viability of individual cells, thereby enabling deeper insights into the potential mechanisms of cell death. In ORP analysis with PCV-1, we identified 0.3 as the cutoff point for assessing whether adding a given drug will cause apparent cytotoxicity, which greatly expands the probe's applicability. To the best of our knowledge, PCV-1 is the first probe to allow visualizing cell death and cell injury under super-resolution imaging, and our proposed analytical assay using it paves the way for quantifying cell viability at the single-cell level.
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This review considers the main properties of fish gelatin that determine its use in food technologies. A comparative analysis of the amino acid composition of gelatin from cold-water and warm-water fish species, in comparison with gelatin from mammals, which is traditionally used in the food industry, is presented. Fish gelatin is characterized by a reduced content of proline and hydroxyproline which are responsible for the formation of collagen-like triple helices. For this reason, fish gelatin gels are less durable and have lower gelation and melting temperatures than mammalian gelatin. These properties impose significant restrictions on the use of fish gelatin in the technology of gelled food as an alternative to porcine and bovine gelatin. This problem can be solved by modifying the functional characteristics of fish gelatin by adding natural ionic polysaccharides, which, under certain conditions, are capable of forming polyelectrolyte complexes with gelatin, creating additional nodes in the spatial network of the gel.
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The purpose of this review article is to examine the method of making gelatin, the characteristics of gelatin from the results of research that has been carried out in Indonesia and the benefits of fish gelatin. Based on a review of various articles and other literature, it can be concluded that fish bone gelatin can be extracted by the acid method. The production of fishbone gelatin consists of 4 stages, the preparation of raw materials includes removal of non-collagen components from raw materials, conversion of collagen to gelatin, purification of gelatin by filtering and finally drying and powdering. Fishbone gelatin can be applied to both the food and non-food industries.
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Gelatin is a product of hydrolysis of collagen protein from animals that are partially processed. Gelatin used in food and non food industries. Gelatin is produced when many import of raw skins and bones of pigs and cows. Goat skins potential as a raw material substitution that still doubt its halal. Process production of gelatin determine the properties of gelatin. The objectives of this research were to determine amino acid profile, group of functional and molecular weight distribution of gelatin made from goat skins which was produced through a process of acid. The skin of male Bligon goat, 1.5 to 2.5 year old was used as raw materials. Process production of gelatin was using acid type acetic acid (CH 3 COOH 0.5 M) (v/v) as curing material. The experimental design applied in this study and commercial gelatin was used as control. The results showed that gelatin produced from goat skin through the process of acid had properties identical with commercial gelatin. It can be concluded that the gelatin has the potential substitute product of commercial gelatin. Keywords : collagen, gelatin, goat skin, curing, acid process
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Amino Acid Analysis
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Physical properties of shark gelatin were examined during gel formation and postgelation in comparison with pig gelatin. Samples with various concentrations and pH values were evaluated by breaking strength, dynamic viscoelasticity, and dynamic light scattering. Sol−gel and gel−sol transition temperatures for shark gelatin were remarkably lower than those for pig gelatin. Shark gelatin gel shows a narrower pH range to form a stable gel compared with pig gelatin. Melting enthalpy of shark gelatin gel was greater than that of pig gelatin gel, and G' of shark gelatin gel changed more extensively with rising temperature in comparison with pig gelatin gel. It is concluded that shark gelatin has different characteristics from pig gelatin not only for gel characteristics but also for the solution property. Keywords: Gelatin; rheology; viscoelasticity; shark; gel
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The combined modification (phthalation followed esterification) of gelatin and the effect of esterification on the isoelectric point of phthalated gelatin were studied. The experimental results showed.1. The isoeleetric point of the parent alkali-processed gelatin is 4.57. 2. After the carboxy groups of gelatin were esterified with ethanol, the isoeleetric point of esterified gelatin may be raised up to 5.78 even to 9.60 depending on the degree of esterification. 3. The isoeleetric point of esterified and then phthalated gelatin (PEA gelatin) was 0.48 pH unit higher than that of phthalated gelatin (PA gelatin). 4. The coagulating property of PA and PEA gelatin differs greatly.
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Fish gelatin, partially hydrolyzed or denatured collagen, has received attention as the alternative of mammalian and avian gelatin. It can be extracted from the by-products rich in collagen such as fish skin, bone, scale, or the skin of some invertebrate, and so on. Composition and properties of fish gelatin can be governed by the sources of raw materials. Processing parameters such as pretreatment, extraction temperature, bleaching, drying, and so on influence the chemical and functional properties of gelatin. Generally, fish gelatin exhibited lower functional properties than those from land animals, thereby limiting the applications of fish gelatin. Several approaches have been therefore developed to improve the properties of fish gelatin via modification using chemical or enzymatic processes. Therefore, fish gelatin can be applied more widely in several industries.
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Gelatin, a well-known biocompatible polymer, is a derivative of collagen obtained by the heat denaturation of collagen. The properties of gelatin vary depending on the sources and extraction techniques. Gelatin is remarkably known for its special gel-forming ability, which makes it a suitable material for exploring the fundamental functional features in colloid studies. Gelatin is widely explored by the scientific community because of its unique characteristics and is highly demanded by various industries such as food, pharmaceutical, cosmetic, photographic, etc. This chapter focuses on the various sources, extraction, and properties of gelatin and provide an insight into gelatin in recent applications.
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