After CNTs functionalizing and dispersion, the CNTs suspension was sterilized by autoclaving (Napco TM Autoclave VA, USA) for 20 min at 118°C.[30 33 34 (link)] The alginate solution was prepared by dissolving alginate powder (Sigma) in sodium chloride (0.9%) as a 1.5% (w/v) solution. To eliminate the contaminant, the alginate solution was passed through a filter with a pore size of 0.2 μm. 0.1 and 1 μg/ml of aseptic CNTs suspension was mixed with 2 ml of sterile alginate solution and the mixture was then stirred for 24 h using a magnetic stirrer.[23 (link)]
Alginate powder
Manufactured by Merck Group
Sourced in United States, Germany
Alginate powder is a natural polysaccharide derived from brown seaweed. It functions as a thickening, stabilizing, and gelling agent in various applications, including food, pharmaceutical, and industrial settings.
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Lab products found in correlation
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Dopamine, by Merck Group (1 mentions)
Freezone 2.5 liter benchtop freeze dry system, by Labconco (1 mentions)
Tensor 27, by Bruker (1 mentions)
Silver flakes, by Merck Group (1 mentions)
Alginate, by Merck Group (1 mentions)
Gfp huvec, by American Type Culture Collection (1 mentions)
Agarose powder, by Biowest (1 mentions)
Celltracker red, by Thermo Fisher Scientific (1 mentions)
Xylazine, by Merck Group (1 mentions)
Eosin stain, by Merck Group (1 mentions)
Hematoxylin stain, by Merck Group (1 mentions)
Acetic acid, by Merck Group (1 mentions)
Ethanol, by Merck Group (1 mentions)
Hydrochloric acid (hcl), by Merck Group (1 mentions)
Sodium chloride (nacl), by Merck Group (1 mentions)
Paraffin, by Merck Group (1 mentions)
Formalin, by Merck Group (1 mentions)
Xylene, by Merck Group (1 mentions)
Bovine fibrinogen, by Merck Group (1 mentions)
8 protocols using alginate powder
1
Preparing Sterile CNT-Alginate Suspensions
2
Alginate Solution Preparation Protocol
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A 1.2% (w/v) alginate solution was made by dissolving 0.360 g of alginate powder (Sigma) slowly in 30 ml of phosphate buffered saline containing a magnetic stirrer. The solution was then autoclaved at 120 °C for 20 minutes.
3
Synthesis and Characterization of Alginate-Catechol Conjugate
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ALG-C was synthesized via EDC-NHS chemistry. Briefly, 100 mg of alginate powder (Sigma-Aldrich, Oakville, ON, USA) was dissolved in 10 mL of distilled water. Then, 143 mg of 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC·HCl), 86 mg of N-hydroxysuccinimide (NHS), and 130 mg of dopamine (Sigma-Aldrich) were added to the alginate solution. The mixture was reacted overnight at room temperature. Then, unreacted EDC, NHS, and dopamine were dialyzed using dialysis membrane (MWCO: 12 K Da, Spectra/Por®) against distilled water for 3 days. The distilled water was changed three times each day. After lyophilization at −50 °C for 3 days (FreeZone 2.5 Liter Benchtop Freeze Dry System, Labconco), the alginate-catechol conjugate was collected. The degree of substitution was analyzed by 1H NMR spectroscopy (Avance III 400 FT-NMR, Bruker). Briefly, 5 mg of ALG-C was dissolved in 1 mL of deuterium oxide (D2O, Sigma-Aldrich). Then, ALG-C solution was loaded in Wilmad® NMR tube and analyzed. In addition, the synthesis of ALG-C was further characterized by Attenuated Total Reflection-Fourier Transformation Infrared (ATR-FTIR, TENSOR27, Bruker). Briefly, the lyophilized ALG-C was scanned 32 times with a resolution of 8 cm−1 and measured from wavenumbers ranging from 650 to 4000 cm−1. The ALG-C film was also fabricated via the above process using CaCl2.
4
Alginate-Methylcellulose Bioprinted Scaffolds
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Alginate powder (Sigma) was sterilized by steam sterilization. 3% alginate solution was prepared in PBS and 9% methylcellulose (MC, Fluka), sterilized by gamma irradiation or scCO2 treatment was added and allowed to swell for additional 2 h at RT. After final stirring, the polysaccharide blend was plotted with the 3D Bio-Scaffold Printer (BioScaffolder 2.1, Gesim, Germany). Cross-linking of the final construct was achieved by soaking in a 100 mM CaCl2 solution for 30 min.
5
Inductive Wireless Charging via Conductive Ink
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The polymer solution was prepared by dissolving 5% Alginate powder (Sigma-Aldrich) in deionized water followed by centrifuging for 2 min at a rate of about 7155 × g to remove air bubbles. The conductive ink was prepared by adding silver flakes (with an average diameter of 10 µm, Sigma-Aldrich) and ethanol into the Alginate solution in the weight ratio of 4:6:1. The square resistivity of the ink film was measured with four-point probing equipment (ST2558B-F01, Suzhou Jingge Electronic co., Ltd.). Through wireless power transmission, an alternating magnetic field was generated by an electromagnetic coil with a power of 1600 W. An LED connected to the conductive silver structures was used to demonstrate the inductive currents generated by the alternating magnetic field.
6
Tissue Engineering Scaffold Fabrication
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Type I collagen derived from rat tail was prepared followed by previously described protocols and stored at 4°C. CaCl2 and sodium hydroxide (NaOH) powder were purchased from Aladdin (Shanghai, China). Agarose powder with low melting temperature (87–89°C) was bought from Biowest (Spain). Alginate solution with a viscosity of 2000 mPa·S was prepared by dissolving Alginate powder (Sigma, United Kingdom) into the cell culture medium with 10% (w/v) fetal bovine serum and 1% (w/v) antibiotic/antimycotic. 0.1 mol/L NaOH solution was prepared by dissolving NaOH powder into phosphate buffer saline (PBS) at room temperature. 3% (w/v) CaCl2 solution was prepared by dissolving CaCl2 powder into tris-buffered saline (TBS) at room temperature. 2% (w/v) flat agarose hydrogel was prepared by casting the boiling agarose solution with 3% (w/v) CaCl2 into a petri dish at room temperature. Before the printing process, the pH of the collagen solution was adjusted to 7.2-7.6. GFP expressing human umbilical vein endothelial cells (GFP-HUVEC; ATCC, USA) and embryonic rat cardiomyocytes (H9C2; ATCC, USA) labeled with red cell tracker (Invitrogen, cat. no. C34552) were cultured in Dulbecco’s Modified Eagle Medium (DMEM, Thermo) for printing cell-laden constructs.
7
Histological Tissue Preparation Protocol
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The following chemicals including ethanol (Merck Company, Germany), paraffin, eosin stain, hematoxylin stain, xylene (Merck Company, Germany), formalin (Merck Company, Germany), acetic acid (Merck Company, Germany), ketamine HCl, xylazine, alginate powder, NaCl (Merck Company, Germany), distilled water, and HCl (Merck Company, Germany) were purchased.
8
Fibrinogen-Alginate Biomaterial Ink Preparation
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The fibrinogen-based biomaterial ink was prepared by dissolving bovine fibrinogen (65–85% protein, Merck, Darmstadt, Germany) in deionized water to have a final concentration of 30 mg/mL. The solution was mixed using a magnetic stirrer at room temperature for 30 min, then alginate powder (medium viscosity, Merck) was added to obtain a final concentration of 8% (w/v). The biomaterial ink was semicrosslinked by mixing the fibrinogen-alginate solution with 100 mM CaCl2 at volumetric ratios of 25:9 (v/v). The sterile formulation was obtained using alginate powder prepared according to a previously described protocol [21 (link)], sterile fibrinogen, and CaCl2 and by performing all procedures under a laminar hood with sterile tools. The semicrosslinked biomaterial ink was centrifuged at 2500 RPM for 5 min to remove air bubbles.
The pH of the final formulation was measured using pH indicator strips.
The pH of the final formulation was measured using pH indicator strips.
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