Calcium Alginate Hydrogel Research Articles

Elasticity-Modulated Microbeads for Classification of Floating Normal and Cancer Cells Using Confining Microchannels.

Engineered microbeads have a wide range of applications in cancer research including identification, characterization, and sorting of cancer cells. In particular, the microbead-based cancer identification techniques are mainly based on the known genetic or biochemical biomarkers; and detection specificity is yet to be improved. On the other hand, it has been discovered that biomechanical properties of cancer cells such as cell-body elasticity can be considered as cancer biomarkers. Here, we report a straightforward microfluidic classification scheme for floating/dissociated normal and cancer epithelial cells using a confining microchannel device together with calcium-alginate hydrogel microbeads. The hydrogel microbeads are generated based on the microfluidic emulsion process, with characterization on the process parameters (e.g., liquid driving pressure and cross-linking duration) in order to specify the resultant bead diameter and elasticity. These engineered microbeads are first mixed with a cell mixture of dissociated human nasopharyngeal epithelial cells (NP460) and nasopharyngeal carcinoma cells (NPC43). The cell elasticity can then be reflected from the locations of captured cells in the device. Experiments further demonstrate that the cell classification has a success rate of >95%. Furthermore, we performed the microbead-based cell classification on a whole blood sample containing floating human breast epithelial cells (MCF-10A) and breast cancer epithelial cells (MDA-MB-231) with a success rate of >75%, revealing its directly applicability to identification of circulating tumor cells in human blood. Together, this research demonstrates a new application of engineered hydrogel microbeads for classification of cells based on their mechanical properties.

Facile tuning of the mechanical properties of a biocompatible soft material

Herein, we introduce a method to locally modify the mechanical properties of a soft, biocompatible material through the exploitation of the effects induced by the presence of a local temperature gradient. In our hypotheses, this induces a concentration gradient in an aqueous sodium alginate solution containing calcium carbonate particles confined within a microfluidic channel. The concentration gradient is then fixed by forming a stable calcium alginate hydrogel. The process responsible for the hydrogel formation is initiated by diffusing an acidic oil solution through a permeable membrane in a 2-layer microfluidic device, thus reducing the pH and freeing calcium ions. We characterize the gradient of mechanical properties using atomic force microscopy nanoindentation measurements for a variety of material compositions and thermal conditions. Significantly, our novel approach enables the creation of steep gradients in mechanical properties (typically between 10–100 kPa/mm) on small scales, which will be of significant use in a range of tissue engineering and cell mechanosensing studies.

Adsorption and Electrochemical Detection of Bovine Serum Albumin Imprinted Calcium Alginate Hydrogel Membrane

In this paper, bovine serum albumin (BSA)-imprinted calcium alginate (CaAlg) hydrogel membrane was prepared using BSA as a template, sodium alginate (NaAlg) as a functional monomer, and CaCl2 as a cross-linker. The thickness of the CaAlg membrane was controlled by a glass rod enlaced with brass wires (the diameter was 0.1, 0.2, 0.3, 0.4, and 0.5 mm). The swelling properties of the CaAlg membranes prepared with different contents of NaAlg were researched. Circular dichroism indicated that the conformation of BSA did not change during the preparing and eluting process. The thinner the CaAlg hydrogel membrane was, the larger the adsorption capacity and the higher the imprinting efficiency of the CaAlg. The maximum adsorption capacity of molecularly imprinted polymer (MIP) and non-imprinted CaAlg hydrogel membrane (NIP) was 38.6 mg·g−1 and 9.2 mg·g−1, respectively, with an imprinting efficiency of 4.2. The MIP was loaded on the electrode to monitor the selective adsorption of BSA by voltammetry curve.

CaAlg hydrogel containing bone morphogenetic protein 4-enhanced adipose-derived stem cells combined with osteochondral mosaicplasty facilitated the repair of large osteochondral defects.

Cartilage repair presents a challenge to clinicians and researchers. A more effective procedure that can produce hyaline-like cartilage is needed for articular cartilage repair. Mosaic osteochondral grafts for large osteochondral defects often show poor integration between the grafts and the surrounding normal cartilage, leading to defective cracks filled with fibrous tissue instead of hyaline-like cartilage. In the present study, we aimed to repair the defective cracks with a calcium alginate (CaAlg) hydrogel containing bone morphogenetic protein 4 (BMP4)-enhanced adipose-derived stem cells (ADSCs). ADSCs were transduced with BMP4 (B-ADSCs). The expression of BMP4 and type II collagen was confirmed using an enzyme-linked immunosorbent assay (ELISA). Swine models of large cartilage defects of the knee were constructed and received one of the four treatments: mosaicplasty only, mosaicplasty with the CaAlg hydrogel, mosaicplasty with the CaAlg hydrogel containing ADSCs, or mosaicplasty with the CaAlg hydrogel containing B-ADSCs injected into the defective cracks. Outcomes were evaluated at 12 and 24weeks after surgery. The in vitro study showed that the osteogenic and chondrogenic activities of the B-ADSCs were enhanced compared with those of the control. In vivo, in the group that received mosaicplasty-containing B-ADSCs, osteochondral tissue was completely integrated with an intact surface. Additionally, the histological scores of the mosaicplasty-containing B-ADSCs group were significantly higher than those of the other groups. Biomechanical examination confirmed that the neocartilage possessed properties similar to those of normal cartilage. Mosaicplasty and hydrogel containing B-ADSCs promoted the repair of large cartilage defects by regenerating hyaline cartilage and repairing dead spaces between osteochondral grafts and donor-site defects, thus improving the feasibility and success rate of one-stage complete repair surgery for large osteochondral defects. This proposed method provides a novel and effective means for the repair of large articular osteochondral defects.

Development of alginate hydrogel/gum Arabic/gelatin based composite capsules and their application as oral delivery carriers for antioxidant.

We have designed microcapsules-immobilized composite capsules and evaluated the oral delivery efficacy. The composite capsules were developed by encapsulating Perinereis aibuhitensis extract (PaE), a model substance possessing antioxidant activity, loaded gum Arabic/gelatin microcapsules in calcium alginate (CA) hydrogel (PaE:CA/GA/GE-CCs). In vitro antioxidant assay showed the obtained composite capsules were able to protect PaE from gastric acid, since O2- scavenging rate of encapsulated PaE was about 1.8 folds as that of free PaE after 5 h incubation in simulated gastrointestinal fluid. Moreover, in vivo study showed that after the treatment of oral administration for 30 days, the mice of PaE:CA/GA/GE-CCs group suffered significantly lower oxidative stress level than those of other groups, illustrated as higher SOD and catalase activity, as well as lower malondialdehyde content in liver cells. The results demonstrated the composite capsules could concentrate PaE in small intestine, and enhance the absorption efficiency and in vivo efficacy.

Immobilization of enological pectinase in calcium alginate hydrogels: A potential biocatalyst for winemaking

A biocatalyst was obtained by immobilizing an enological commercial pectinase within calcium alginate hydrogels using an entrapment technique, and its catalytic activity was evaluated during different storage conditions. Hydrogel beads were stored at 4 °C in three different ways: (i) wet, in citrate buffer solution (pH 3.8); (ii) dehydrated by using a vacuum stove; and (iii) freeze-dried. Biocatalyst surface and their internal morphology were characterized by Scanning Electron Microscopy and a good enzyme distribution throughout alginate matrix was observed. Fourier Transform Infrared Spectroscopy results confirmed the presence of absorption bands associated with amino groups present in enzymes. Immobilization procedure did not modify the optimal pH and temperature (pH = 4.0 and 20 °C) for pectinase activity, comparing to free enzyme. Entrapped pectinase showed activity until six reaction cycles with 40% residual activity. Storage stability studies demonstrated that wet entrapped pectinase retained its initial enzymatic activity up to 11 weeks, whereas that lyophilized hydrogels retained its original activity after 8 months of storage. These results suggest that immobilized pectinase may be successfully exploited in various industrial applications, with special concern in grape juice clarification process. Thus, the turbidity of grape must decreased significantly using the immobilized pectinase during 150 min at 20 °C. This biocatalyst could be easily removed after clarification process and it can be reused, minimizing production economic costs in wine industry.

Removal of heat stable salts from industrial lean methyldiethanolamine using magnetic alginate/iron oxide hydrogel composite

Heat Stable Salts (HSS) are undesirable contaminates that are generated due to the degradation of the amine solvent. This study deals with the development of a new process for the removal of HSS from amine solvent using magnetic calcium alginate hydrogel composites. The magnetic calcium alginate hydrogel composites was prepared by encapsulating freshly prepared maghemite nanoparticles (γ-Fe2O3) synthesized using a conventional precipitation technique in the calcium alginate beads. The developed magnetic hydrogel composites (CA-Fe2O3) characterized by XRD, SEM and FTIR showed that γ-Fe2O3 are retained homogeneously in the composites matrix. Kinetic and equilibrium batch adsorption experiments revealed that maximum HSS removal of 29.24% was achieved in 240 min with 3.0 g of onto CA-Fe2O3 composites at 25 °C and sorption efficiency increased with an increase in temperature. Adsorption kinetic study was performed using three kinetic models and the pseudo-second order kinetics very well correlated with the experimental data; demonstrating the chemisorption and endothermic nature of sorption. The reusability experiments showed no significant loss in absorption capability even after eight cycles. Additionally, the superparamagnetic behavior showcased by the magnetic composites allowed easy separation of the composites from lean MDEA using an external magnet. Therefore, such magnetic alginate–Fe2O3 hydrogel composites can work as a simple and cost-effective adsorbent for the reclamation of industrial lean MDEA, with the additional advantages of good efficiency, eco-friendliness and being easy to fabricate and recover.

Preparation of composite fabric loaded with mi-croencapsulated plant extracts and its inhibitory effect on lipase

PurposeThis study aims to elaborate on the microencapsulation of the plant extract (PE, from Camellia sinensis leaf, clover flower and cocoa flower) and the preparation of a slow-release lining fabric loading the PE microcapsule.Design/methodology/approachPE was microencapsulated into polyvinyl alcohol (PVA) shells through interfacial polymerization. The morphology, thermal stability, slow-release property and drug loading ratio of the PVA/PE microcapsules were characterized to ensure the availability in coating finishing. To find the optimum parameters, the composite fabrics were prepared from non-woven fabrics coated by calcium alginate hydrogel, which glued mass fractions of microcapsules and dried in different ways. To evaluate the effectiveness, a lipase enzyme activity test was conducted.FindingsUnder optimal conditions, the PVA/PE microcapsules with smooth surface have an average particle size of 14.5 um, and they are expected to reach a loading ratio of 38.5 per cent while remaining stable under 220°C. Given a microcapsule of 4 per cent (of the mass), the composite fabric has a good hand feeling, being prepared through calcium chloride coating. It is shown that the inhibition ratios of the microcapsules and composite fabrics on lipase are 31.3 and 21.0 per cent, respectively.Research limitations/implicationsThe composite fabric could be prepared through the other finishing methods such as padding and printing. In addition, the release mechanism of the composite could be studied.Practical implicationsThis study provided a simple and effective way to prolong the duration of PE. This way was conductive to protect environmental sensitive PEs from being destroyed in compositing processes.Originality/valuePreparing composite fabrics for transdermal delivery system was novel and other kind of plant extracts could be used in this way.

Direct writing alginate bioink inside pre-polymers of hydrogels to create patterned vascular networks

We describe a strategy to fabricate a hydrogel-based microvascular construct by direct writing alginate bioink inside the viscous pre-polymer of hydrogels, which acts as a support bath. As the print needle translates through the polymers, the extruded alginate instantaneously forms calcium alginate hydrogel (Ca-Alg) templates deposited within the bath. This phase change allows the formed templates to be anchored within the pre-polymers, while maintaining their structure. After the printing process, the pre-polymers are solidified to form a mechanically robust hydrogel. Finally, a hydrogel construct with embedded microchannels is generated by liquefying and removing the Ca-Alg templates. Using this method, not only the alginate ink alone can be directly printed within the engineered constructs, but also the size and shape of the formed microchannels are controllable. Furthermore, a confluent endothelial layer for the generation of vascular networks can be constructed by adhering and proliferating endothelial cells on the channel linings. This strategy demonstrates a promising technique for rapid construction of in vitro vasculatures, which would provide a versatile platform for a wide array of applications such as tissue engineering, organ-on-a-chip and drug screening.

A Cocktail-Inspired Male Birth Control Strategy with Physical/Chemical Dual Contraceptive Effects and Remote Self-Cleared Properties.

Inspired by cocktails, we designed a medium term (2-20 weeks) male contraceptive strategy. Through a sequential injection process of four reagents (calcium alginate hydrogel, PEG-Au nanoparticles (PEG-AuNps), EDTA, and PEG-AuNps), physical clogging of the vas deferens and chemical inhibition of the sperm motility were realized simultaneously. The contraceptive period could be directly preset by adjusting the injection ratio of each reagent. More interesting, the embolism area could be readily dredged through a short-time noninvasive near-infrared irradiation. The present study offered an effective and reversible manner to fill the gap of current medium-term contraceptive strategy. In addition, the proposed in vivo pipeline plugging technology, with a flexible noninvasive self-cleared characteristic, might also provide a convenient and reliable strategy for some other biomedical engineering researches.

Algisyl® Injections: An Innovative Strategy for Patients with Advanced Heart Failure

The remodelling of the heart that occurs after myocardial infarction independently contributes to heart failure (HF) progression. In a rapidly growing patient population with chronic HF, new, safe, and effective therapeutic strategies are needed. Recently, injectable biomaterial-based therapies have been gaining interest, especially for patients post myocardial infarction who have had complications due to advanced HF. One such intervention, based on Algisyl® (LoneStar Heart Inc., Irvine, California, USA) injections, has been examined in the recent prospective, randomised, controlled AUGMENT-HF clinical trial. This paper briefly presents characteristics of Algisyl, an injectable calcium alginate hydrogel; describes a minimally invasive myoplasty procedure for the insertion of Algisyl implants; and provides a concise overview of the design, methods, and results of the AUGMENT-HF study. This paper focusses on the promising findings that Algisyl, in addition to standard medical therapy, was more effective than the standard medical therapy alone for providing sustained 6-month and 1-year benefits in exercise capacity and symptomatic improvement among patients with advanced HF. In addition, this report discusses some implications and challenges relevant to the AUGMENT-HF trial, and addresses the importance of a blind study design for further studies in this field. Moreover, this paper highlights future perspectives for examining the Algisyl implants, aimed at functional clinical outcomes, quality of life, and safety issues, prior to a possible implementation of this strategy into clinical practice.

Cobalt-mediated multi-functional dressings promote bacteria-infected wound healing

Wound dressings with multiple functions are required to meet the complexity of the wound healing process. The multifunctionality often leads to an increase in the complexity and difficulty in dressing preparation. To surmount this problem, we used a facile preparation and fabrication process to fabricate a multi-functional dressing by integrating four widely accessible materials: plain gauze, sodium alginate (SA), Ca2+ and Co2+. Firstly, mixed Ca2+/Co2+ ion solutions with different concentration were applied to gauzes. After drying, SA solution was added to ionized gauze and Co2+-Ca2+/Gauze/SA (Ion-GSA) composite dressings were formed easily. In vitro results showed that all Ion-GSA dressings exhibited strong mechanical properties, uniform dispersion and sustained release of Ca2+ and Co2+, and the ability to retain moisture and absorb wound exudate. Besides the above advantages, dressings prepared with 0.25 g/L Co2+ and 4 g/L Ca2+ (Co2+0.25-Ca2+4 GSA composite dressings) exhibited the best overall effect for inducing a hypoxia-like response, and favorable cytocompatibility, hemostatic property and antibacterial activity. In vivo wound healing assays revealed that Co2+0.25-Ca2+4 GSA composite dressings inhibited bacterial growth, increased local Hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF), transforming growth factor-β1 (TGF-β1) protein expression, and accelerated full-thickness skin wound healing in mouse bacterial-infected wound model. The quick healing wounds had improved angiogenesis, macrophages regulation, re-epithelialization and dense collagen deposition. Collectively, our results indicated that Co2+0.25-Ca2+4 GSA composite dressings promote wound healing. Statement of SignificanceWound dressings with integrated functionalities are required to meet complex clinical requirements. However, there is often a trade-off between reducing preparation complexity and increasing the multifunctionality of the dressing’s properties. In this study, we prepared multifunctional composite dressings by a facile preparation process using widely accessible materials. The composite dressings possessed the mechanical strength of gauze, had the effective wound exudate absorption, moisture maintenance and hemostatic property capacity of calcium alginate hydrogels, and had the hypoxia-like induction and the antimicrobial effects of Co2+. These functions all together promote bacteria-infected wound healing. Thus, we believed that the composite dressings can be widely applied in skin wound repair duo to their facile preparation method and good therapeutic effect.

Microfluidic synthesis of highly spherical calcium alginate hydrogels based on external gelation using an emulsion reactant

Alginate-based hydrogels are widely used in the biomedical and chemical fields, and their size and shape are significant to their applications like drug delivery and cell encapsulation. Here, we report a microfluidic external gelation process using an on-chip calcium chloride (CaCl2) emulsion reactant for producing highly spherical calcium alginate (Ca-alginate) hydrogel particles. The microfluidic channels were two serial cross-junctions fabricated on quartz glass. Monodisperse sodium alginate (Na-alginate) droplets with diameters greater or smaller than the opening (ranging from 176 μm to 225 μm) with a coefficient of variation (CV) less than 3% were successfully generated at the upstream cross-junction; they then reacted with the CaCl2 emulsion at the downstream cross-junction, forming Ca-alginate hydrogels. The effects of the fraction of the aqueous phase in the reactant CaCl2 emulsion and the flow rates of continuous and emulsion phases on the roundness of the obtained hydrogels were studied. By optimizing the parameters above, monodisperse spherical hydrogel particles were obtained with diameters ranging from 147 μm to 176 μm with CVs around 5%. The synthesis of magneto-responsive hydrogels with asymmetric Fe3O4 coatings was also demonstrated.

PH-responsive calcium alginate hydrogel laden with protamine nanoparticles and hyaluronan oligosaccharide promotes diabetic wound healing by enhancing angiogenesis and antibacterial activity.

Diabetic wounds as chronic wounds represent a severe, persistent complication of diabetes and, in the most extreme cases, can lead to amputation. Two critical and unfavorable factors affecting diabetic wound healing are sustained bacterial-induced chronic inflammation and disrupted vascularization. In this paper, we presented a novel, pH-responsive calcium alginate hydrogel laden with protamine nanoparticles and hyaluronan oligosaccharides, and explored its potential for accelerating diabetic wound healing. A thorough investigation indicated that the drug- and nanoparticle-loaded hydrogel demonstrated strong bactericidal behavior mediated by protamine nanoparticles and reduced bacterial-induced chronic inflammation at the wound site. Furthermore, it accelerated the wound-healing process by promoting angiogenesis in skin wounds with the hyaluronan oligosaccharide-mediated enhanced expression of vascular endothelial growth factor.

Alginate-Based Hydrogel Beads as a Biocompatible and Efficient Adsorbent for Dye Removal from Aqueous Solutions.

In this study, sodium alginate was employed as a starting material for preparing two kinds of biocompatible adsorbents, including calcium alginate hydrogel beads and magnetic hydrogel beads. Fourier transform infrared spectroscopy, X-ray diffraction pattern, and scanning electron microscopy/energy-dispersive X-ray techniques were used to characterize the prepared adsorbents. The performance of the prepared adsorbents for the removal of methyl violet from aqueous solution was studied in detail. Both kinetics and equilibrium aspects of methyl violet adsorption were investigated, and the obtained equilibrium and kinetics data were described with various adsorption models. The effects of initial dye concentration, adsorbent dosage, and temperature on adsorption performance were investigated. Thermodynamic parameters of adsorption were obtained as well.

In-situ determination of time-dependent alginate-hydrogel formation by mechanical texture analysis

Although alginate-based hydrogels are widely used, determining their mechanical properties and explaining their ability to form a hydrogel in situ remains a challenge. A new method of external gelation using calcium chloride (CaCl2) is proposed to quantify the formation of alginate hydrogel and its mechanical properties as well as to observe its temporal behaviour. To this end, small inhomogeneous alginate beads were formed, which comprised a shell and a core. The bead’s shell consisted of calcium alginate hydrogel, while its core was sodium alginate sol. The beads were mechanically tested with a texture analyser to provide a detailed description of the structure. Different alginate samples could be clearly identified, and the M(mannuron)/G(guluron) acid ratios could be extrapolated quickly and easily from the test results if calibrated by NMR. This enables a fast and cost-effective characterisation of different alginates with regard to their composition and gel formation properties.

Injectable composite of calcium alginate hydrogel and Adipose-derived Stem Cells to remediate bone defect

To investigate the feasibility of injectable composite of calcium alginate hydrogel and adipose-derived stem cells(ASCs) to remediate bone defect, the experiment was carried out. ASCs from New Zealand rabbit cultured in vitro were mixed with 1.5% sodium alginate and calcium gluconate, preparing the calcium alginate hydrogel with cell density 5x109/L. The cell growth curve was used to determine the biocompatibility of ASCs and hydrogels. The mixture was injected into the radius defect area of the New Zealand rabbits(group A); the calcium alginate hydrogel without cells was injected into the same area of other rabbits(group B). All rabbits were examined with radiodetection and bone mineral density(BMD) in the 12 postoperative weeks. Radiological assessment showed that 12 weeks after injection, the bone formation was obvious in experimental group A. In group B, a small number of new bone appeared at both ends, with the defect still visible. In group C, no bone formation was found. It also indicated that from 4 to 12 weeks post-surgery, the average BMD value of group A was higher than group B and group C(P < 0.01). Percutaneous injection of calcium alginate hydrogel and ASCs composites can help to remediate bone defect in animals.

Relative Humidity and Temperature Sensing Characteristics of an Optical Fiber Probe Based on Hollow-Core Fiber and Calcium Alginate Hydrogel Film

Hydrogel is a kind of functional cross-linked polymer material with 3-D network structure. When hydrogel is exposed to water environment, a mass of hydrophobic groups and hydrophilic residues existing in the cross linked netlike nano-structure is willing to combine with water molecules and expand, respectively. In this paper, a novel optical fiber probe based on hollow-core fiber and calcium alginate (CaAlg) hydrogel film is proposed and experimentally demonstrated. The experimental results of relative humidity (RH) and temperature sensing characteristics show that the proposed probe has a binomial response to RH in the range of 20%~90% RH and a linear response to temperature with the sensitivity of 121pm/°C. CaAlg is considered as an effective and potential as the humidity-sensitive material to optical fiber RH sensors.

Entrapment and growth of Chlamydomonas reinhardtii in biocompatible silica hydrogels.

In this work, we aimed at improved viability and growth of the microalga Chlamydomonas reinhardtii in transparent silica hydrogels based on low-ethanol, low-sodium and low-propylamine synthesis. Investigation into replacement of conventional base KOH by buffers dipotassium phosphate and tris(hydroxymethyl)aminomethane along with increased precursor concentrations yielded an aqueous synthesis route which provided a gelation within 10 min, absorptions below 0.1 and elastic moduli of 0.04-4.23 kPa. The abrasion resistance enhanced by 41% compared to calcium alginate hydrogels and increased to 70-85% residual material on addition of chitosan. Entrapment of microalgae in low-sodium and low-propylamine silica hydrogels maintained the PSII quantum yield above 0.3 and growth rates of 0.23 ± 0.01 d-1, similarly to cells entrapped in calcium alginate. These promising results pave the way for the entrapment of sensitive, photosynthetically active and growing cells for in robust biotechnological applications.

Test standard and output force characteristics of the electroresponsive biomimetic actuator based on calcium alginate hydrogel

Since Sodium Alginate (SA) hydrogel has disadvantages of poor controllability and low mechanical strength, it is difficult to satisfy the practical requirements when used alone. In this paper, with CaCl2 as the crosslinker, Calcium Alginate Hydrogel (CAH) was prepared as electric actuating membrane via ionic crosslinking. The non-metallic electrode membrane was fabricated from SA blended with Multi-walled Carbon Nanotube (MWCNT). Further, two electrode membranes were stuck on both sides of the actuating membrane to assemble a biodegradable Electroresponsive Biomimetic Actuator (ERBA). Moreover, an experimental platform was built for testing the output force of the ERBA cross-linked with different CaCl2 concentrations, and experimental test standards were developed according to preparation conditions and applied voltage. Then output force density was utilized to evaluate the output force characteristics. Based on in-depth analysis of the ERBA actuating mechanism, the influencing mechanisms of elastic modulus, specific capacitance and CaCl2 concentration on output force performance of the ERBA was illustrated respectively. The experimental results showed that suitable amount of the membrane solutions was 40 ml for preparing the ERBA, and its optimal test voltage was 3 V DC. When CaCl2 concentration was 0.9 g L−1, the ERBA was best cross-linked, where its elastic modulus was large and ion migration rate was fast, which caused the maximum output force density of 26.534 mN g−1. But as the CaCl2 concentration increased, the actuating membrane was cross-linked excessively, which resulted in blocked internal ion channel, small ion migration speed and reduced output force.