https://www.nature.com/articles/s41598-025-12075-7
The article presents a milli-fluidic device and electronic extrusion system for fabricating alginate/carbon nanotube microfibers using calcium chloride as a crosslink agent. The device was designed, simulated, and prototyped, and successfully applied the mixture of alginate and carbon nanotube to generate microfibers in different concentrations and stepper motor speeds. The flow extrusion pump was used to pump fluid flow to the milli-fluidic device and crosslink calcium chloride to form sodium alginate-CNT microfibers. The microfibers were fabricated and characterized using techniques like FE-SEM, FTIR, Raman Spectroscopy, and XRD. The results showed the material’s amorphous crystallinity and the composite nature of the fibbers, consisting of alginate as the matrix and CNTs as the reinforcing, conductive filler. The invented technology successfully generated microfibers with sizes ranging from 60 to 100 μm and I-V measurements were tested. The study demonstrates the potential of the developed system for fabricating conductive microfibers with properties relevant to nerve tissue engineering. However, further biological validation is required to confirm their suitability for nerve repair applications. The findings have significant implications for the design of scaffolds in regenerative nerve therapies. These findings support the potential of the fabricated microfibers as conductive scaffolds for nerve tissue engineering, although further biological validation is required.
The need for advanced nerve tissue engineering solutions is evident in the face of the persistent challenges associated with peripheral nerve injuries. Traditional strategies for nerve repair have evolved to include more sophisticated approaches like nerve- guided catheters, which combine scaffolding and cells, and have shown potential in promoting significant functional recovery. Additionally, non-surgical treatments such as laser irradiation and traditional Chinese medicine have been explored for their neurogenerative effects. These advancements highlight the urgency and potential in developing new materials and techniques for nerve repair and regeneration1.
Carbon nanotubes have garnered attention in nerve regeneration research due to their unique properties. However, current strategies for peripheral nerve regeneration, including those involving CNTs, face challenges. Despite advancements in surgical reconstruction and postoperative rehabilitation, patients often experience lingering motor and sensory deficits. This underscores the need for novel therapeutic strategies and technologies that can effectively promote nerve regeneration and enhance functional outcomes2. The fabrication of the alginate and CNT has garnered significant attention in recent years due to highly stable nanomaterials that exhibit remarkable mechanical and electrical characteristics, including exceptional tensile strength and flexibility. Fabrication technology has witnessed remarkable advancements over the past few decades, enabling precise control and manipulation of materials at the micro- and nanoscale3. Milli-fluidics has emerged as a powerful platform for various biological studies due to its ability to mimic physiological conditions and its high-throughput capabilities4. This technique offers precise control over the diameter and cross- sectional shape of microfibers through the manipulation of microchannel device size, geometry, and flow rate ratio5. Milli-fluidics enables the continuous production of fibers with highly tunable geometries and mechanical properties6. Traditional microfiber fabrication methods, such as electrospinning, often face challenges in achieving consistent fiber diameter, morphology, and uniform distribution of nanomaterials7. Additionally, electrospinning may not be suitable for incorporating hydrophilic materials like alginate due to their tendency to clog the spinneret8. Numerous techniques have been suggested to produce carbon nanotube (CNT) microfibers. The three most established methods are: the direct growth of CNTs through chemical vapor deposition9,10,11; spinning CNT fibers from nanotube forests12,13, wet spinning from solutions containing CNTs and surfactants14,15.To address the limitations of traditional methods, this thesis proposes milli- fluidic approach for fabricating alginate and carbon nanotube (CNT) microfibers. This method utilizes a milli-fluidic device to control the flow of alginate and CNTs, enabling the production of microfibers with consistent diameter, morphology, and uniform CNT dispersion16. The milli-fluidic environment facilitates uniform dispersion of CNTs within the alginate matrix, preventing clumping and clogging. This uniform dispersion is crucial for achieving the desired properties of the microfibers, such as electrical conductivity and mechanical strength17.
The proposed milli-fluidic approach offers several advantages over traditional methods such as precise control over microfiber diameter and morphology, the microchannel dimensions and flow rates can be precisely controlled to achieve consistent microfiber characteristics18. This level of control is difficult to achieve with traditional methods, such as electrospinning, which can be susceptible to variations in fiber diameter and morphology due to factors such as environmental conditions and spinneret clogging19. The second advantage is the uniform CNT dispersion, the milli-fluidic environment facilitates uniform dispersion of CNTs within the alginate matrix, preventing agglomeration and clogging20. This uniform dispersion is essential for achieving the desired properties of microfibers, such as mechanical strength and electrical conductivity. Traditional methods, such as electrospinning, can be prone to CNT agglomeration, which can negatively impact the properties of the microfibers21,22,23,24. The third advantage is the scalability, the milli-fluidic device can be easily scaled up for large-scale production of microfibers. This scalability is important for commercial applications of microfibers, such as in tissue engineering and drug delivery. Traditional methods, such as electrospinning, can be time-consuming and difficult to scale up for large-scale production24– 26. An electronic extrusion system integrated with the milli- fluidic device to enable real-time monitoring and control of the fabrication process was proposed in27. This integration allows for precise control of fluid flow rates, chemical concentrations, and crosslinking durations, ensuring consistent microfiber properties28. The fabrication of microfibers using milli-fluidic technology enables precise control over the diameter and morphology of the microfibers, leading to consistent and tunable properties. This level of control is challenging to achieve with traditional methods29,30,31.
The alginate and CNT microfibers hold promise for various applications due to their unique properties, including biocompatibility, biodegradability, and electrical conductivity. These properties make them suitable for a range of applications in tissue engineering, biomedical devices, and environmental remediation32. In the field of tissue engineering, alginate and CNT microfibers can serve as scaffolds for nerve regeneration. These microfibers mimic the extracellular matrix (ECM), providing a supportive environment for neural cell growth and differentiation33. The biocompatibility and biodegradability of alginate allow it to be gradually degraded as the nerve tissue regenerates, while the electrical conductivity of CNTs promotes nerve cell adhesion and outgrowth34. Alginate and CNT microfibers can also be used for drug delivery applications, as they can be loaded with drugs and then implanted into the body, where they can slowly release the drugs in a controlled manner. This targeted drug delivery approach can reduce systemic side effects and improve drug efficacy35,36,37,38. In biomedical devices, alginate and CNT microfibers can be used to create implants, biosensors, and diagnostic tools39,40,41,42,43. For example, alginate and CNT microfibers can be used to fabricate neural electrodes for recording neural activity or stimulating nerve cells. The electrical conductivity of alginate and CNTs allows for efficient transfer of electrical signals between the electrodes and the nervous system, and CNT microfibers can also be used in environmental remediation applications44. In this study, the application proposed for the fabricated microfibers of alginate and carbon nanotube is nerve regulation because the carbon nanotube microfibers (CNT) will be used to repair damaged nervous system tissue. Damaged nerve tissues are reconnected by biopolymer, but these biopolymers are non-electrically conductive, so we need CNTs to connect the signals between the tissues, as CNTs are conductive materials. The electrical conductivity of CNTs can promote nerve cell adhesion and neurite outgrowth45. The mechanical strength and porosity of the CNT microfibers provide support for nerve cells and promote their growth and differentiation46,47,48,49. Hence, the researchers are continuing to explore the potential of CNT microfibers for nerve regulation applications, and there is great promise for these materials in the future. This study represents a comprehensive exploration of the development process, covering the design, simulation, fabrication, and characterization of the fabricated CNT microfibers. It delves into the intricacies of milli-fluidic technology, nanomaterials, and electronic control systems, with the goal of contributing to the advancement of materials science and related fields. The milli-fluidic device was designed using Google SketchUp software, simulated using COMSOL Multiphysics software, and then printed using a 3D printer and PDMS replication, while the electronic extrusion system was designed to control the flow of fluids and the addition of chemicals to the milli-fluidic device, including the crosslinking agent (CaCl2) to obtain the microfibers of alginate and CNT, which are then applied to the nerve regulation application.
Damaged neuron-cell communication and nerve degeneration are the prevalent manifestations of nerve injury50. The human brain consists of a vast assembly of meticulously arranged neurons, each executing special functions and facilitating communication with the body via the nervous system. Neurons, the primary nerve cells in the brain, transmit information through a combination of electrical and chemical signals throughout the nervous system51. Tissues in the central nervous system (CNS) are unique because they cannot be restored. Regeneration of neural tissue is hampered by its dynamic nature, difficulties in repairing the blood-brain barrier, and disruption of secondary tissue52. Therefore, it’s important that any method aiming to repair the central nervous system mainly focuses on copying injured axons, fixing nerve signals, and growing new neurons. When the central nervous system is damaged, it often causes astrocytosis, where numerous astrocytes are produced. At the same time, these reactive astrocytes can harm both nerve and non-nerve cells53. This makes it impossible to produce axons in neurons and restore nervous tissue in a balanced manner. Injuries during orthopedic surgery can also harm nerves. The reaction of nerves to severe injuries can lead to nerve function failure. First, problems with nerve function may manifest as coordination issues or difficulty recalling names. These issues can worsen if many neurons break down. Depending on the type of nervous system damage, restoring the activity of damaged nerves or treating damage from neurodegenerative disorders is a significant concern in the field of biomedicine54. Neuronal development involves creating, fixing, or substituting weak neurons and nerve structures. This includes generating new axons, synapses, neurons, glia, and myelin sheaths. For materials applied to nerve tissue, they need to conduct electricity to support cell production, break down naturally, and have bioactivity for delivering growth factors. Neurodegenerative disorders typically occur because the neural network is damaged, leading to brain disorders55. The combination of sodium alginate CNT has not been widely explored in the context of nerve regulation48,49,52. The advantages over existing materials, such as enhanced nerve cell adhesion and neurite outgrowth, along with the alginate and CNT microfibers exhibiting improved electrical conductivity compared to traditional biomaterials, promote nerve cell adhesion and neurite outgrowth. This enhanced conductivity is crucial for interfacing with the nervous system and facilitating nerve regeneration17,56,57.
In this study, we proposed a new method to produce CNT microfibres in which alginate and carbon nanotubes were combined and mixed to fabricate sodium alginate-CNT microfibres. We use the milli-fluidic device platform and the electronic extrusion system to carefully control the flow rate and concentration of the sodium alginate-CNT solutions. This allows us to produce uniform sodium alginate-CNT microfibres, which we can then use in nerve regulation system applications to repair nervous tissue damage. Damage nervous tissue that is reconnected by biopolymers, but these biopolymers are non-electrically conductive (insulators), which means they are not providing a high surface area for volume, and it’s not small enough. Therefore, we need the intervention of nanomaterials, specifically carbon nanotubes. These nano-conductive materials are used because, first, they are electrically conductive to provide signals to the cells, have a high surface area over a high surface volume, and are small enough to facilitate the diffusion of nutrients and growth factors. This allows for the restoration of electrical conductivity signals between the damaged nervous tissues. Secondly, the nano size of the nanotubes allows for the diffusion of nutrients and growth factors due to their porous nature, and thirdly, they do not integrate well with the cells. However, when combined with alginate, an FDA-approved non-conductive material, the biocompatibility of carbon nanotubes increased.
The study employs a structured research method to establish, design, develop, and characterize experiments, as illustrated in flow chart Fig. 1. It begins with an overview of the experimental framework, followed by detailed steps in the creation and assessment of the milli-fluidic devices and microfibres. The methodology includes the design of a 3D mold for millifluidic devices and the fabrication of a specialized millifluidic extrusion head. It also includes the simulation processes for milli-fluidics and the making of PDMS milli-fluidic devices, which shows how precise and efficient these steps need to be. We also discuss the development of an extrusion pump, which plays a pivotal role in the controlled delivery and application of materials in the experiments. A key focus is on the generation of microfibres composed of alginate and carbon nanotubes, which are essential for the study’s objective. The characterisation process employed a variety of analytical techniques, including FTIR, Raman spectroscopy, XRD, SEM, and I-V measurement. Each of these methods provides critical insights into the composition, structure, and properties of the synthesised microfibres. Lastly, the chapter details an experiment to apply these sodium alginate-CNT microfibers in simulating nerve fibers, which is central to understanding their potential applications in biomedical contexts. This approach exemplifies a comprehensive methodology that combines advanced fabrication techniques with detailed analytical assessments, aiming to contribute significantly to the field of nerve tissue regeneration research.
The flow chart of the methodology of the study.