2D carbon network arranged into high-order 3D nanotube arrays on a flexible microelectrode: integration into electrochemical microbiosensor devices for cancer detection

The research focuses on advancing electrochemical biosensors for cancer detection using novel carbon-based nanomaterials. Existing low-dimensional nanocarbon materials, while possessing desirable properties for catalysis and biosensing, suffer from restacking and aggregation, limiting their effectiveness. This study addresses this limitation by developing a strategy to arrange nanocarbon materials into higher-order 3D architectures, enhancing their surface area and stability. The researchers utilize ionic liquids (ILs) as precursors for synthesizing a novel 3D nanotube array (NTA) structure composed of a mesoporous 2D N, B, and P codoped carbon network (NBP-CNW). ILs offer advantages such as low vapor pressure, high thermal stability, and tunable chemical structures, enabling the creation of carbon nanomaterials with controlled morphology and properties. The chosen synthesis method uses a 3D ZnO nanorod array (NRA) template grown on activated carbon fibers (CFs), providing a controlled structure for the NBP-CNW growth. This approach allows for the creation of a highly porous, heteroatom-doped carbon structure with enhanced electrochemical activity, crucial for efficient biosensing applications. The overall goal is to create a highly sensitive, stable, and biocompatible microelectrode for detecting H2O2, a significant cancer biomarker, offering a potential breakthrough in early cancer diagnosis and treatment monitoring.

The introduction references the extensive research on various carbon allotropes (graphene quantum dots, nanodiamonds, carbon nanotubes, graphene, and carbon networks) and their applications in catalysis, energy storage, biosensing, and biomedicine. It highlights the challenges associated with the aggregation and restacking of low-dimensional nanocarbons, reducing their effective surface area and stability. The use of template methods to create higher-order architectures from these materials is emphasized as a crucial advancement to overcome these challenges. The review also touches upon the use of different carbon precursors (glucose, polydopamine, and heteroatom polymers) and acknowledges the ongoing challenge of synthesizing carbon nanomaterials with precise control over their nanostructure, chemical and electronic properties, and multifunctionality. The use of ionic liquids (ILs) as a novel precursor is introduced, highlighting their unique properties and potential for creating tunable, heteroatom-doped carbon structures.

The methodology section describes the synthesis of the NBP-CNW-NTAs/CF microelectrode in detail. Activated carbon fibers (CFs) serve as the base material, onto which ZnO nanorod arrays (NRAs) are grown via electrodeposition. A mixture of two ionic liquids, [VEIM]BF4 and [OMIM]PF6 (4:1 volume ratio), is then coated onto the ZnO-NRAs. The coated structure is subjected to high-temperature treatment (750°C under Ar atmosphere) to transform the IL layer into a N, B, and P co-doped carbon layer. The ZnO-NRAs template is subsequently removed using 0.1 M HCl solution. The resultant NBP-CNW-NTAs/CF microelectrode is characterized using SEM and TEM, revealing its unique 3D hierarchical porous structure. The electrochemical properties of the microelectrode are evaluated. The fabrication of both a microfluidic chip and an implantable probe incorporating the NBP-CNW-NTAs/CF microelectrode is detailed. Hepatoma (HepG2), cervical cancer (HeLa), and breast cancer (MCF-7) cells are cultured and used for evaluating the biosensor performance. The cytotoxicity of the microelectrode is also assessed using a CCK-8 assay. Surgically resected human breast tumor specimens are used to evaluate the performance of the implantable probe in distinguishing tumor tissue from normal tissue. The electrochemical measurements are performed using a three-electrode system with the NBP-CNW-NTAs/CF microelectrode as the working electrode.

The synthesized NBP-CNW-NTAs exhibit a unique 3D hierarchical porous structure, as confirmed by SEM and TEM analysis. This structure contributes to a large electrochemically active surface area (ECSA), abundant active sites, and efficient charge transport pathways, leading to enhanced electrocatalytic activity and stability. Heteroatom doping (N, B, P) modifies the electronic conductivity and wettability of the carbon material. The NBP-CNW-NTAs/CF microelectrode demonstrates excellent sensing performance towards H2O2, with a low detection limit (500 nM), a wide linear dynamic range (up to 19.52 μM), and high sensitivity (61.8 μA cm−2 mM−1). The microfluidic chip incorporating this microelectrode enables real-time monitoring of H2O2 secretion from different cancer cell lines (HepG2, HeLa, MCF-7), with and without radiotherapy, allowing for differentiation of cancer cell types and evaluation of therapeutic efficacy. The implantable probe successfully differentiates tumor tissues from normal tissues in surgically resected human breast specimens. These findings demonstrate the potential of this novel electrochemical microbiosensor for accurate and minimally invasive cancer diagnostics.

The results demonstrate the successful fabrication of a high-performance electrochemical microbiosensor based on a novel 3D hierarchical porous carbon nanostructure. The superior sensing performance stems from the synergistic effects of the unique morphology, high surface area, abundant active sites, and efficient charge transfer characteristics of the NBP-CNW-NTAs. The ability to distinguish between different cancer cell lines and assess radiotherapy efficacy highlights the potential for personalized cancer treatment. The successful in situ detection of cancer tissues in human specimens underscores the clinical applicability of the implantable probe for minimally invasive cancer diagnosis. The study contributes to the development of advanced electrochemical biosensors for early cancer detection and improved therapeutic monitoring. The use of ionic liquids as precursors offers a versatile and sustainable approach to designing advanced carbon-based nanomaterials for various biomedical applications.

This study successfully synthesized a novel 3D hierarchical porous NBP-CNW-NTAs/CF microelectrode exhibiting excellent electrochemical sensing performance for H2O2. Its integration into a microfluidic chip and an implantable probe provides a powerful tool for real-time cancer cell monitoring and in situ tissue diagnosis. The results demonstrate a significant advancement in minimally invasive cancer detection and personalized therapy monitoring. Future research could explore the application of this technology to other cancer types and investigate the optimization of the sensor design for enhanced sensitivity and selectivity.

The study primarily focuses on three types of cancer cells (HepG2, HeLa, MCF-7) and breast cancer tissue. Further research is needed to validate the performance of this sensor with a broader range of cancer types and tissue samples. While the implantable probe demonstrates promise, its long-term stability and biocompatibility in vivo require additional investigation. The influence of other interfering species present in biological samples on the sensor's performance also needs further investigation.