Advances in Electrochemical Nano-Biosensors for Biomedical and Environmental Applications: From Current Work to Future Perspectives.

Modern life quality is strongly supported by the advances made in biosensors, which has been attributed to their crucial and viable contribution in point-of-care (POC) technology developments. POC devices are exploited for the fast tracing of disease progression, rapid analysis of water, and food quality assessment. Blood glucose meters, home pregnancy strips, and COVID-19 rapid tests all represent common examples of successful biosensors. Biosensors can provide great specificity due to the incorporation of selective bio-recognition elements and portability at significantly reduced costs. Electrochemical biosensor platforms are one of the most advantageous of these platforms because they offer many merits, such as being cheap, selective, specific, rapid, and portable. Furthermore, they can be incorporated into smartphones and various analytical approaches in order to increase their sensitivity and many other properties. As a very broad and interdisciplinary area of research and development, biosensors include all disciplines and backgrounds from materials science, chemistry, physics, medicine, microbiology/biology, and engineering. Accordingly, in this state-of-the-art article, historical background alongside the long journey of biosensing construction and development, starting from the Clark oxygen electrode until reaching highly advanced wearable stretchable biosensing devices, are discussed. Consequently, selected examples among the miscellaneous applications of nanobiosensors (such as microbial detection, cancer diagnosis, toxicity analysis, food quality-control assurance, point of care, and health prognosis) are described. Eventually, future perspectives for intelligent biosensor commercialization and exploitation in real-life that is going to be supported by machine learning and artificial intelligence (AI) are stated.

Formation of electroactive biofilms derived by nanostructured anodes surfaces.

Microbial fuel cells (MFCs) have significant interest in the research community due to their ability to generate electricity from biodegradable organic matters. Anode materials and their morphological structures play a crucial role in the formation of electroactive biofilms that enable the direct electron transfer. In this work, modified electrodes with nanomaterials, such as multiwalled carbon nanotubes (MWCNTs), reduced graphene oxide (rGO), Al2O3/rGO or MnO2/MWCNTs nanocomposites were synthesized, characterized and utilized to support the growth of electrochemically active biofilms. The MFC's performance is optimized using anode-respiring strains isolated from biofilm-anode surface, while the adjusted operation is conducted with the consortium of (Enterobacter sp.). Besides the formation of matured biofilm on its surface, MnO2/MWCNTs nanocomposite produced the highest electrical potential outputs (710 mV) combined with the highest power density (372 mW/m2). Thus, a correlation between the anode nanostructured materials and the progression of the electrochemically active biofilms formation is presented, allowing new thoughts for enhancing the MFC's performance for potential applications ranging from wastewater treatment to power sources.

Microbial Electrochemical Systems: Principles, Construction and Biosensing Applications.

Microbial electrochemical systems are a fast emerging technology that use microorganisms to harvest the chemical energy from bioorganic materials to produce electrical power. Due to their flexibility and the wide variety of materials that can be used as a source, these devices show promise for applications in many fields including energy, environment and sensing. Microbial electrochemical systems rely on the integration of microbial cells, bioelectrochemistry, material science and electrochemical technologies to achieve effective conversion of the chemical energy stored in organic materials into electrical power. Therefore, the interaction between microorganisms and electrodes and their operation at physiological important potentials are critical for their development. This article provides an overview of the principles and applications of microbial electrochemical systems, their development status and potential for implementation in the biosensing field. It also provides a discussion of the recent developments in the selection of electrode materials to improve electron transfer using nanomaterials along with challenges for achieving practical implementation, and examples of applications in the biosensing field.

Electrochemical Impedance Spectroscopy (EIS): Principles, Construction, and Biosensing Applications.

Electrochemical impedance spectroscopy (EIS) is a powerful technique used for the analysis of interfacial properties related to bio-recognition events occurring at the electrode surface, such as antibody-antigen recognition, substrate-enzyme interaction, or whole cell capturing. Thus, EIS could be exploited in several important biomedical diagnosis and environmental applications. However, the EIS is one of the most complex electrochemical methods, therefore, this review introduced the basic concepts and the theoretical background of the impedimetric technique along with the state of the art of the impedimetric biosensors and the impact of nanomaterials on the EIS performance. The use of nanomaterials such as nanoparticles, nanotubes, nanowires, and nanocomposites provided catalytic activity, enhanced sensing elements immobilization, promoted faster electron transfer, and increased reliability and accuracy of the reported EIS sensors. Thus, the EIS was used for the effective quantitative and qualitative detections of pathogens, DNA, cancer-associated biomarkers, etc. Through this review article, intensive literature review is provided to highlight the impact of nanomaterials on enhancing the analytical features of impedimetric biosensors.

Microbial Sensing and Removal of Heavy Metals: Bioelectrochemical Detection and Removal of Chromium(VI) and Cadmium(II).

The presence of inorganic pollutants such as Cadmium(II) and Chromium(VI) could destroy our environment and ecosystem. To overcome this problem, much attention was directed to microbial technology, whereas some microorganisms could resist the toxic effects and decrease pollutants concentration while the microbial viability is sustained. Therefore, we built up a complementary strategy to study the biofilm formation of isolated strains under the stress of heavy metals. As target resistive organisms, Rhizobium-MAP7 and Rhodotorula ALT72 were identified. However, Pontoea agglumerans strains were exploited as the susceptible organism to the heavy metal exposure. Among the methods of sensing and analysis, bioelectrochemical measurements showed the most effective tools to study the susceptibility and resistivity to the heavy metals. The tested Rhizobium strain showed higher ability of removal of heavy metals and more resistive to metals ions since its cell viability was not strongly inhibited by the toxic metal ions over various concentrations. On the other hand, electrochemically active biofilm exhibited higher bioelectrochemical signals in presence of heavy metals ions. So by using the two strains, especially Rhizobium-MAP7, the detection and removal of heavy metals Cr(VI) and Cd(II) is highly supported and recommended.

Point-of-Care Diagnostics of COVID-19: From Current Work to Future Perspectives.

Coronaviruses have received global concern since 2003, when an outbreak caused by SARS-CoV emerged in China. Later on, in 2012, the Middle-East respiratory syndrome spread in Saudi Arabia, caused by MERS-CoV. Currently, the global crisis is caused by the pandemic SARS-CoV-2, which belongs to the same lineage of SARS-CoV. In response to the urgent need of diagnostic tools, several lab-based and biosensing techniques have been proposed so far. Five main areas have been individuated and discussed in terms of their strengths and weaknesses. The cell-culture detection and the microneutralization tests are still considered highly reliable methods. The genetic screening, featuring the well-established Real-time polymerase chain reaction (RT-PCR), represents the gold standard for virus detection in nasopharyngeal swabs. On the other side, immunoassays were developed, either by screening/antigen recognition of IgM/IgG or by detecting the whole virus, in blood and sera. Next, proteomic mass-spectrometry (MS)-based methodologies have also been proposed for the analysis of swab samples. Finally, virus-biosensing devices were efficiently designed. Both electrochemical immunosensors and eye-based technologies have been described, showing detection times lower than 10 min after swab introduction. Alternative to swab-based techniques, lateral flow point-of-care immunoassays are already commercially available for the analysis of blood samples. Such biosensing devices hold the advantage of being portable for on-site testing in hospitals, airports, and hotspots, virtually without any sample treatment or complicated lab precautions.

Biological Insights of Fluoroaryl-2,2'-Bichalcophene Compounds on Multi-Drug Resistant Staphylococcus aureus.

Resistance of bacteria to multiple antibiotics is a significant health problem; hence, to continually respond to this challenge, different antibacterial agents must be constantly discovered. In this work, fluoroaryl-2,2'-bichalcophene derivatives were chemically synthesized and their biological activities were evaluated against Staphylococcus aureus (S. aureus). The impact of the investigated bichalcophene derivatives was studied on the ultrastructural level via scanning electron microscopy (SEM), molecular level via sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) method and on the biofilm inhibition via the electrochemical biosensors. Arylbichalcophenes' antibacterial activity against S. aureus was affected by the presence and location of fluorine atoms. The fluorobithiophene derivative MA-1156 displayed the best minimum inhibitory concentration (MIC) value of 16 µM among the tested fluoroarylbichalcophenes. Over a period of seven days, S. aureus did not develop any resistance against the tested fluoroarylbichalcophenes at higher concentrations. The impact of fluoroarylbichalcophenes was strong on S. aureus protein pattern showing high degrees of polymorphism. SEM micrographs of S. aureus cells treated with fluoroarylbichalcophenes displayed smaller cell-sizes, fewer numbers, arranged in a linear form and some of them were damaged when compared to the untreated cells. The bioelectrochemical measurements demonstrated the strong sensitivity of S. aureus cells to the tested fluoroarylbichalcophenes and an antibiofilm agent. Eventually, these fluoroarylbichalcophene compounds especially the MA-1156 could be recommended as effective antibacterial agents.

Multifunctional Nanotechnology-Enabled Sensors for Rapid Capture and Detection of Pathogens.

Nanomaterial-based sensing approaches that incorporate different types of nanoparticles (NPs) and nanostructures in conjunction with natural or synthetic receptors as molecular recognition elements provide opportunities for the design of sensitive and selective assays for rapid detection of contaminants. This review summarizes recent advancements over the past ten years in the development of nanotechnology-enabled sensors and systems for capture and detection of pathogens. The most common types of nanostructures and NPs, their modification with receptor molecules and integration to produce viable sensing systems with biorecognition, amplification and signal readout are discussed. Examples of all-in-one systems that combine multifunctional properties for capture, separation, inactivation and detection are also provided. Current trends in the development of low-cost instrumentation for rapid assessment of food contamination are discussed as well as challenges for practical implementation and directions for future research.

Mediated bioelectrochemical system for biosensing the cell viability of Staphylococcus aureus.

Staphylococcus aureus is one of the most dangerous human pathogens and is the cause of numerous illnesses ranging from moderate skin infections to life-threatening diseases. Despite advances made in identifying microorganisms, rapid detection methods for the viability of bacteria are still missing. Here, we report a rapid electrochemical assay for cell viability combining the use of double redox mediators and multiwall carbon nanotubes-screen printed electrodes (MWCNTs-SPE), ferricyanide (FCN) and 2,6-dichlorophenolindophenol (DCIP), which served as electron shuttle to enable the bacterial-electrode communications. The current originating from the metabolically active cells was recorded for probing the activity of the intracellular redox centers. Blocking of the respiratory chain pathways with electron transfer inhibitors demonstrated the involvement of the electron transport chain in the reaction. A good correlation between the number of the metabolically active cells and the current was obtained. The proposed assay has been exploited for monitoring cell proliferation of S. aureus during the growth. The sensitivity of the detection method reached 0.1 OD600. Therefore, the technique described is promising for estimating the cell number, measuring the cell viability, and probing intracellular redox center(s).

Regulation of Candida albicans Interaction with Macrophages through the Activation of HOG Pathway by Genistein.

The severity of infections caused by Candida albicans, the most common opportunistic human fungal pathogen, needs rapid and effective antifungal treatments. One of the effective ways is to control the virulence factors of the pathogen. Therefore, the current study examined the effects of genistein, a natural isoflavone present in soybeans, on C. albicans. The genistein-treated C. albicans cells were then exposed to macrophages. Although no inhibition effect on the growth rates of C. albicans was noted an enhancement of the immune response to macrophages has been observed, indicated by phagocytosis and release of cytokines TNF-α and IL-10. The effect of genistein on the enhanced phagocytosis can be mimicked by the fungicides fludioxonil or iprodione, which inhibit the histidine kinase Cos1p and lead to activation of HOG pathway. The western blot results showed a clear phosphorylation of Hog1p in the wild type strain of C. albicans after incubation with genistein. In addition, effects of genistein on the phosphorylation of Hog1p in the histidine kinase mutants Δcos1 and Δsln1 were also observed. Our results thus indicate a new bio-activity of genistein on C. albicans by activation of the HOG pathway of the human pathogen C. albicans.

Bacteriophage-based nano-biosensors for the fast impedimetric determination of pathogens in food samples.

The early and rapid detection of pathogenic microorganisms is of critical importance in addressing serious public health issues. Here, a new bacteriophage-based nano-biosensor was constructed and the electrochemical impedimetric method was fully optimized and applied for the quantitative detection of Escherichia coli O157:H7 in food samples. The impact of using a nanocomposite consisting of gold nanoparticles (AuNPs), multi-walled carbon nanotubes (MWCNTs), and tungsten oxide nanostructures (WO3) on the electrochemical performance of disposable screen printed electrodes was identified using the cyclic voltammetry and electrochemical impedance spectroscopy. The use nanomaterials enabled high capturing sensitivity against the targeting bacterial host cells with the limit of detection of 3.0 CFU/ml. Moreover, selectivity of the covalently immobilized active phage was tested against several non-targeting bacterial strains, where a high specificity was achieved. Thus, the targeting foodborne pathogen was successfully detected in food samples with high specificity, and the sensor provided an excellent recovery rate ranging from 90.0 to 108%. Accordingly, the newly developed phage-biosensor is recommended as a disposable label-free impedimetric biosensor for the quick and real-time monitoring of food quality.

Non-enzymatic disposable electrochemical sensors based on CuO/Co3O4@MWCNTs nanocomposite modified screen-printed electrode for the direct determination of urea.

A new electrochemical impedimetric sensor for direct detection of urea was designed and fabricated using nanostructured screen-printed electrodes (SPEs) modified with CuO/Co3O4 @MWCNTs. A facile and simple hydrothermal method was achieved for the chemical synthesis of the CuO/Co3O4 nanocomposite followed by the integration of MWCNTs to be the final platform of the urea sensor. A full physical and chemical characterization for the prepared nanomaterials were performed including Fourier-transform infrared spectroscopy (FTIR), Raman spectroscopy, X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), contact angle, scanning electron microscope (SEM) and transmission electron microscopy (TEM). Additionally, cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to study the electrochemical properties the modified electrodes with the nanomaterials at different composition ratios of the CuO/Co3O4 or MWCNTs. The impedimetric measurements were optimized to reach a picomolar sensitivity and high selectivity for urea detection. From the calibration curve, the linear concentration range of 10-12-10-2 M was obtained with the regression coefficient (R2) of 0.9961 and lower detection limit of 0.223 pM (S/N = 5). The proposed sensor has been used for urea analysis in real samples. Thus, the newly developed non-enzymatic sensor represents a considerable advancement in the field for urea detection, owing to the simplicity, portability, and low cost-sensor fabrication.

Designing and fabrication of electrochemical nano-biosensor for the fast detection of SARS-CoV-2-RNA.

SARS-CoV-2 caused a global panic among populations. Rapid diagnostic procedures for the virus are crucial for disease control. Thus, the designed signature probe from a highly conserved region of the virus was chemically immobilized onto the nanostructured-AuNPs/WO3-screen printed electrodes. Different concentrations of the matched oligonucleotides were spiked to test the specificity of the hybridization affinity whereas the electrochemical impedance spectroscopy was used for tracking the electrochemical performance. After a full assay optimization, limits of detection and quantification were calculated based on linear regression and were valued at 298 and 994 fM, respectively. Further, the high performance of the fabricated RNA-sensor chips was confirmed after testing the interference status in the presence of the mismatched oligos in one nucleotide and completely one. Worthy to mention that the single-stranded matched oligos can be hybridized to the immobilized probe in 5 min at room temperature. The designed disposable sensor chips are capable of detecting the virus genome directly. Therefore, the chips are a rapid tool for SARS-CoV-2 detection.

Ferrocene tagged primary antibody generates electrochemical signal: An electrochemical immunosensing platform for the monitoring of vitamin D deficiency in clinical samples.

In this paper, a new kind of ultrasensitive and low-cost electrochemical immunosensing probe was designed to monitor vitamin D deficiency using 25(OH)D3 as a clinical biomarker. Ferrocene carbaldehyde conjugated on Ab-25(OH)D3 antibodies was used as an electrochemical probe for generating signals. The graphene nanoribbon-modified electrode (GNRs) was used to immobilize the (Ab-25(OH)D3-Fc) conjugate. The high electron transferability, greater surface area, and effective biocompatibility of GNRs enabled the capture of the greater number of primary antibodies (Ab-25(OH)D3). The developed probe was structurally and morphologically characterized. The step-wise modification was investigated by electrochemical techniques. The direct electrochemistry of ferrocene enabled 25(OH)D3 biomarker detection with excellent sensitivity. The reduction in peak current was proportional to the concentrations of 25(OH)D3 in the range of 1-100 ng mL-1 with a 0.1 ng mL-1 limit of detection. The probe was tested in terms of reproducibility, repeatability, and stability. Finally, the developed immunosensing probe was applied in serum samples for 25(OH)D3 quantification, and no significant difference was noticed in the assay results when compared with the standard chemiluminescent immunoassay (CLIA) method. The developed detection strategy has a wider scope for future potential clinical diagnostics applications.

Double-antibody-based nano-biosensing system for the onsite monitoring of SARS-CoV-2 variants.

The fast and reliable diagnosis of COVID-19 is the foremost priority for promoting public health interventions. Therefore, double-antibody-based immunobiosensor chips were designed, constructed, and exploited for clinical diagnosis. Gold nanoparticles/tungsten oxide/carbon nanotubes (AuNPs/WO3/CNTs) were used as the active working sensor surface to support the chemical immobilization of a mixture of SARS-CoV-2 antibodies (anti-RBD-S and anti-RBD-S-anti-Llama monoclonal antibodies). The morphology and chemical functionalization of the fabricated disposable immunochips was characterized using scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). After full assay optimization, the immunobiosensor showed a high sensitivity to detect SARS-CoV-2-S protein with limits of detection and quantification of 1.8 and 5.6 pg/mL, respectively. On the other hand, for the SARS-CoV-2 whole virus particle analysis, the detection and quantification limits were determined to be 5.7 and 17 pg/mL, respectively. The biosensor showed a highly selective response toward SARS-CoV-2, even in the presence of influenza, nontargeting human coronaviruses, and Middle East respiratory syndrome coronavirus (MERS-CoV). The immunochips exhibited distinct responses toward the variants of concern: B.1>C.36.3>Omicron> Delta> Alpha coronavirus variants. For biosensor validation, twenty-nine clinical specimens were analyzed, and the impedimetric responses were positively detected for two Delta samples, eighteen Omicron samples, and six B.1-type samples in addition to three negative samples. Eventually, the immunobiosensor was fabricated in the form of ready-to-use chips capable of sensitive detection of virus variants, especially variants of concern (VOC) and interest, in a specimen within 15 min. The chips provided instantaneous detection with the direct application of clinical samples and are considered a point-of-care device that could be used in public places and hot spots.

Selective determination of nitrite in water and food samples using zirconium oxide (ZrO2)@MWCNTs modified screen printed electrode.

Nitrite ions are being used in different forms as food preservatives acting as flavor enhancers or coloring agents for food products. However, continuous ingestion of nitrite may have severe health implications due to its mutagenic and carcinogenic effects. Thus, this study constructed an electrochemical assay using disposable nano-sensor chip ZrO2@MWCNTs screen printed electrodes (SPE) for the rapid, selective, and sensitive determination of nitrite in food and water samples. As a sensing platform, the use of nanomaterials, including metal oxide nanostructures and carbon nanotubes, exhibited a superior electrocatalytic activity and conductivity. Morphological, structural, and electrochemical analyses were performed using electron microscopy (SEM and TEM), Fourier-transform infrared (FTIR) spectroscopy, electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV) and chronoamperometry (CA). Accordingly, a wide dynamic linear range (5.0 µM to 100 µM) was obtained with a limit of detection of 0.94 µM by the chronoamperometric technique. In addition, the sensor's selectivity was tested when several non-target species were exposed to the sensor chips while no obvious electrochemical signals were generated when the nitrite ions were not present. Eventually, real food and water sample analysis was conducted, and a high recovery was achieved.

Surface grafting of polymeric catheters and stents to prevent biofilm formation of pathogenic bacteria.

BACKGROUND: Tecothane (medical grade of polyurethane) is strongly involved in the fabrication of metallic and polymeric-based medical devices (e.g., catheters and stents) as they can withstand cardiac cycle-related forces without deforming or failing, and they can mimic tissue behavior. The main problem is microbial contamination and formation of pathogenic biofilms on such solid surfaces within the human body. Accordingly, our hypothesis is the coating of tecothane outer surfaces with antibacterial agents through the electro-deposition or chemical grafting of anti-biofilm agents onto the stent and catheter surfaces. RESULTS: Tecothane is grafted with itaconic acid for cross-linking the polyethyleneimine (PEI) as the protective-active layer. Accordingly, the grafting of poly-itaconic acid onto the Tecothane was achieved by three different methods: wet-chemical approach, electro-polymerization, or by using plasma treatment. The successful modifications were verified using Fourier Transform Infrared (FTIR) spectroscopy, grafting percentage calculations, electrochemical, and microscopic monitoring of biofilm formation. The grafting efficiency of itaconic acid was over 3.2% (w/w) at 60 â„ƒ after 6 h of the catheter chemical modification. Bio-electrochemical signals of biofilms have been seriously reduced after chemical modification because of the inhibition of biofilm formation (for both Pseudomonas aeruginosa and Staphylococcus aureus) over a period of 9 days. CONCLUSION: Chemical functionalization of the polyurethane materials with the antimicrobial and anti-biofilm agents led to a significant decrease in the formation of pathogenic biofilms. This promising proof-concept will open the door to explore further surface protection with potential anti-biofilm agents providing better and sustainable productions of stents and catheters biomaterials.

Construction and characterization of nano-oval BaTi0.7Fe0.3O3@NiFe2O4 nanocomposites as an effective platform for the determination of H2O2.

Talented di-phase ferrite/ferroelectric BaTi0.7Fe0.3O3@NiFe2O4 (BFT@NFO) in oval nano-morphology was chemically synthesized using controlled sol-gel processes and calcined at 600 °C. The effects of shielding using NiFe2O4 (NFO) nanoparticles on the microstructure, phase transition, thermal, and relative permittivity of BaTi0.7Fe0.3O3 (BTF) nano-perovskite were systematically explored. X-ray diffraction patterns and Full-Prof software exhibited the forming of the BaTi2Fe4O11 hexagonal phase. TEM and SEM images demonstrated that the coating of BaTi0.7Fe0.3O3 has been successfully controlled with exquisite nano-oval NiFe2O4 shapes. The NFO shielding can significantly promote the thermal stability and the relative permittivity of BFT@NFO pero-magnetic nanocomposites and lowers the Curie temperature. Thermogravimetric and optical analysis were used to test the thermal stability and estimate the effective optical parameters. Magnetic studies showed a decrease in saturation magnetization of NiFe2O4 NPs compared to their bulk system, which is attributed to surface spin disorder. Herein, characterization and the sensitive electrochemical sensor were constructed for the evaluation of peroxide oxidation detection using the chemically adjusted nano-ovals barium titanate-iron@nickel ferrite nanocomposites. Finally, The BFT@NFO exhibited excellent electrochemical properties which can be ascribed to this compound possessing two electrochemical active components and/or the nano-ovals structure of the particles which can further improve the electrochemistry through the possible oxidation states and the synergistic effect. The result advocates that when the BTF is shielded with NFO nanoparticles the thermal, dielectric, and electrochemical properties of nano-oval BaTi0.7Fe0.3O3@NiFe2O4 nanocomposites can be synchronously developed. Thus, the production of ultrasensitive electrochemical nano-systems for the determination of hydrogen peroxide is of extensive significance.

Imprinted Polymers on the Route to Plastibodies for Biomacromolecules (MIPs), Viruses (VIPs), and Cells (CIPs).

Around 30% of the scientific papers published on imprinted polymers describe the recognition of proteins, nucleic acids, viruses, and cells. The straightforward synthesis from only one up to six functional monomers and the simple integration into a sensor are significant advantages as compared with enzymes or antibodies. Furthermore, they can be synthesized against toxic substances and structures of low immunogenicity and allow multi-analyte measurements via multi-template synthesis. The affinity is sufficiently high for protein biomarkers, DNA, viruses, and cells. However, the cross-reactivity of highly abundant proteins is still a challenge.

Bio-electrochemical frameworks governing microbial fuel cell performance: technical bottlenecks and proposed solutions.

Microbial fuel cells (MFCs) are recognized as a future technology with a unique ability to exploit metabolic activities of living microorganisms for simultaneous conversion of chemical energy into electrical energy. This technology holds the promise to offer sustained innovations and continuous development towards many different applications and value-added production that extends beyond electricity generation, such as water desalination, wastewater treatment, heavy metal removal, bio-hydrogen production, volatile fatty acid production and biosensors. Despite these advantages, MFCs still face technical challenges in terms of low power and current density, limiting their use to powering only small-scale devices. Description of some of these challenges and their proposed solutions is demanded if MFCs are applied on a large or commercial scale. On the other hand, the slow oxygen reduction process (ORR) in the cathodic compartment is a major roadblock in the commercialization of fuel cells for energy conversion. Thus, the scope of this review article addresses the main technical challenges of MFC operation and provides different practical approaches based on different attempts reported over the years.