In this report, a facile and label-free electrochemical RNA biosensor is developed by exploiting methylene blue (MB) as an electroactive positive ligand of G-quadruplex. The electrochemical response mechanism of the nucleic acid assay was based on the change in differential pulse voltammetry (DPV) signal of adsorbed MB on the immobilized human telomeric G-quadruplex DNA with a loop that is complementary to the target RNA. Hybridization between synthetic positive control RNA and G-quadruplex DNA probe on the transducer platform rendered a conformational change of G-quadruplex to double-stranded DNA (dsDNA), and increased the redox current of cationic MB π planar ligand at the sensing interface, thereby the electrochemical signal of the MB-adsorbed duplex is proportional to the concentration of target RNA, with SARS-CoV-2 (COVID-19) RNA as the model. Under optimal conditions, the target RNA can be detected in a linear range from 1 zM to 1 μM with a limit of detection (LOD) obtained at 0.59 zM for synthetic target RNA and as low as 1.4 copy number for positive control plasmid. This genosensor exhibited high selectivity towards SARS-CoV-2 RNA over other RNA nucleotides, such as SARS-CoV and MERS-CoV. The electrochemical RNA biosensor showed DPV signal, which was proportional to the 2019-nCoV_N_positive control plasmid from 2 to 200000 copies (R2 = 0.978). A good correlation between the genosensor and qRT-PCR gold standard was attained for the detection of SARS-CoV-2 RNA in terms of viral copy number in clinical samples from upper respiratory specimens.

Most electroanalytical detection schemes for DNA markers require considerable time and effort from expert personnel to thoroughly follow the analysis and obtain reliable outcomes. This work aims to present an electrochemical assay performed inside a small card-based platform powered by microfluidic manipulation, requiring minimal human intervention and consumables. The assay couples a sample/signal dual amplification and DNA-modified magnetic particles for the detection of DNA amplification products. Particularly, the sul1 and sul4 genes involved in the resistance against sulfonamide antibiotics were analyzed. As recognized by the World Health Organization, antimicrobial resistance threatens global public health by hampering medication efficacy against infections. Consequently, analytical methods for the determination of such genes in environmental and clinical matrices are imperative. Herein, the resistance genes were extracted from E. coli cells and amplified using an enzyme-assisted isothermal amplification at 37 °C. The amplification products were analyzed in an easily-produced, low-cost, card-based set-up implementing a microfluidic system, demanding limited manual work and small sample volumes. The target amplicon was thus captured and isolated using versatile DNA-modified magnetic beads injected into the microchannel and exposed to the various reagents in a continuously controlled microfluidic flow. After the optimization of the efficiency of each phase of the assay, the platform achieved limits of detections of 44.2 pmol L-1 for sul1 and 48.5 pmol L-1 for sul4, and was able to detect down to ≥500-fold diluted amplification products of sul1 extracted from E. coli living cells in around 1 h, thus enabling numerous end-point analyses with a single amplification reaction.

Due to the late detection of stomach cancer, this cancer usually causes high mortality. The development of an electrochemical genosensor to measure microRNA 106b (miR-106b), as a gastric cancer biomarker, is the aim of this effort. In this regard, first, 1,3,5-benzenetricarboxylate (BTC) metal-organic frameworks (Zn-BTC MOF) were self-assembled on the glassy carbon electrode and then the probe (ssDNA) was immobilized on it. The morphology Zn-BTC MOF was characterized by SEM, FT-IR, Raman and X-Ray techniques. Zn-BTC MOF as a biosensor substrate has strong interaction with ssDNA. Quantitative measurement of miR-106b was performed by electrochemical impedance spectroscopy (EIS). To perform this measurement, the difference of the charge transfer resistances (ΔRct) of Nyquist plots of the ssDNA probe modified electrode before and after hybridization with miR-106b was obtained and used as an analytical signal. Using the suggested genosensor, it is possible to measure miR-106b in the concentration range of 1.0 fM to 1.0 μM with a detection limit of 0.65 fM under optimal conditions. Moreover, at the genosensor surface, miR-106b can be detected from a non-complementary and a single base mismatch sequence. Also, the genosensor was used to assess miR-106b in a human serum sample and obtained satisfactory results.

The influenza virus is a pervasive pathogen that exhibits increased prevalence during colder seasons, resulting in a significant annual occurrence of infections. Notably, pharmaceutical interventions effective against influenza A strains often exhibit limited efficacy against influenza B variants. Against this backdrop, the need for innovative approaches to accurately and swiftly differentiate and detect influenza B becomes evident. Biosensors play a pivotal role in this detection process, offering rapid, specific, and sensitive identification of the virus, facilitating timely intervention and containment efforts. Oligonucleotide sequences targeting the conserved B/Victoria/2/87 influenza virus NP region were designed. Nasopharyngeal swabs were collected from patients suspected of influenza virus infection, and viral RNA was extracted. RNA quality was assessed through one-step PCR. cDNA synthesis was performed using random hexamers, and real-time PCR quantified the influenza genome. Gold nanoparticles were immobilized on a surface to immobilize the specific DNA probe, and electrochemical hybridization was electrochemically followed. The biosensor exhibited high selectivity and effective distinction of complementary sequences from mismatches and influenza virus cDNA genome. The biosensor successfully detected the influenza B virus genome in real samples. Non-influenza samples yielded no significant hybridization signals. The comparison between the results obtained from the biosensor and real-time PCR revealed full agreement of these methods. The biosensor utilized electrochemical detection of hybridization and proved effective in detecting the influenza B virus genome with high specificity, sensitivity, and selectivity. Comparative analysis with real-time PCR underscored the accuracy and potential applicability of the biosensor in rapid and specific virus detection. This innovative approach holds promise for future diagnostic and epidemiological applications in detecting influenza B virus and other pathogens.

A new electrochemical biosensor based on the sequence of chromosome Y (SRY) has been introduced to determine the gender of the fetus. At first, the DNA probe was designed based on the SRY gene sequence on chromosome Y. Then, a suitable functional group was added to the DNA probe, and it has been immobilized on the surface of the electrode modified with a nanocomposite containing Cu(OH)2 @N-C n-boxes. This substrate causes more DNA probes to connect to the electrode surface by increasing the effective surface area. The presence of the SRY sequence in the DNA sample extracted from blood was detected by the electrochemical signal of the bio-sensor. After optimizing the parameters, the fabricated genosensor showed linear responses in the two concentration ranges containing 0.5 fM to 50 pM and 50 pM to 500 nM. The limit of detection (LOD) for the proposed method was 0.16 fM. The proposed genosensor has been successfully used to determine the gender of the fetus using cell-free fetal DNA (cffDNA) in the blood plasma of several pregnant mothers. This method has advantages such as being simple, portable, accurate, and non-invasive for early determination of the gender of the fetus and early diagnosis of X-linked genetic disorders.

Genosensor-based electrodes mediated with nanoparticles (NPs) have tremendously developed in medical diagnosis. Herein, we report a facile, rapid, low cost and highly sensitive biosensing strategy for early detection of HPV 18 using gold-nanoparticles (AuNPs) deposited on micro-IDEs. This study represents surface charge transduction of micro-interdigitated electrodes (micro-IDE) alumina insulated with silica, independent and mini genosensor modified with colloidal gold NPs (AuNPs), and determination of gene hybridization for early detection of cervical cancer. The surface of AuNPs deposited micro-IDE functionalized with optimized 3-aminopropyl-triethoxysilane (APTES) followed by hybridization with deoxyribonucleic acid (DNA) virus to develop DNA genosensor. The results of ssDNA hybridization with the ssDNA target of human papillomavirus (HPV) 18 have affirmed that micro-IDE functionalized with colloidal AuNPs resulted in the lowest detection at 0.529 aM. Based on coefficient regression, micro-IDE functionalized with AuNPs produces better results in the sensitivity test (R2 = 0.99793) than unfunctionalized micro-IDE.

Acute promyelocytic leukemia (APL) in children is associated with a favorable initial prognosis. However, minimal residual disease (MRD) follow-up remains poorly defined, and relapse cases are concerning due to their recurrent nature. Thus, we report two electrochemical flexible genosensors based on polypyrrole (PPy) and graphene quantum dots (GQDs) for label-free PML-RARα oncogene detection. Atomic force microscopy (AFM), scanning electron microscope (SEM), cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS) were used to characterize the technological biosensor development. M7 and APLB oligonucleotide sequences were used as bioreceptors to detect oncogenic segments on chromosomes 15 and 17, respectively. AFM characterization revealed heterogeneous topographical surfaces with maximum height peaks for sensor layers when tested with positive patient samples. APLB/Genosensor exhibited a percentage change in anode peak current (ΔI) of 423 %. M7/Genosensor exhibited a ΔI of 61.44 % for more concentrated cDNA samples. The described behavior is associated with the biospecific recognition of the proposed biosensors. Limits of detection (LOD) of 0.214 pM and 0.677 pM were obtained for APLB/Genosensor and M7/Genosensor, respectively. The limits of quantification (LOQ) of 0.648 pM and 2.05 pM were estimated for APLB/Genosensor and M7/Genosensor, respectively. The genosensors showed reproducibility with a relative standard deviation of 7.12 % for APLB and 1.18 % for M7 and high repeatability (9.89 % for APLB and 1.51 % for M7). In addition, genetic tools could identify the PML-RARα oncogene in purified samples, plasmids, and clinical specimens from pediatric patients diagnosed with APL with high bioanalytical performance. Therefore, biosensors represent a valuable alternative for the clinical diagnosis of APL and monitoring of MRD with an impact on public health.

The rapid and sensitive detection of food contaminants is becoming increasingly important for timely prevention and treatment of foodborne disease. In this review, we discuss recent developments of electrochemical biosensors as facile, rapid, sensitive, and user-friendly analytical devices and their applications in food safety analysis, owing to the analytical characteristics of electrochemical detection and to advances in the design and production of bioreceptors (antibodies, DNA, aptamers, peptides, molecular imprinted polymers, enzymes, bacteriophages, etc.). They can offer a low limit of detection required for food contaminants such as allergens, pesticides, antibiotic traces, toxins, bacteria, etc. We provide an overview of a broad range of electrochemical biosensing designs and consider future opportunities for this technology in food control.

Fast and effective diagnosis is the first step in monitoring the current coronavirus 2 (CoV-2) pandemic. Herein, we establish a simple and sensitive electrochemical assay using magnetic nanocomposite and DNA sandwich probes to rapidly quantify the CoV-2 nucleocapsid (N) gene down to the 0.37 fM level. This assay uses a pair of specific DNA probes. The capture probe is covalently conjugated to Au-decorated magnetic reduced graphene oxide (AMrGO) nanocomposite for efficiently capturing target RNA. In contrast, the detection probe is linked to peroxidase for signal amplification. The probes target the COV-2 gene, allowing for specific magnetic separation, enzymatic signal amplification, and subsequent generation of voltammetric current with a total assay time of 45 min. The developed biosensor has high selectivity and can discriminate non-specific gene sequences. Synthetic COV-2 N-gene can be detected efficiently in serum and saliva, while 1-bp mismatch gene yielded a low response. The performance of the genosensor was good in an extensive linear range of 5 aM-50 pM. For synthetic N-gene, we achieved the detection limit of 0.37, 0.33, and 0.19 fM in human saliva, urine, and serum. This simple, selective, and sensitive genosensor could have various genetics-based biosensing and diagnostic applications.

Aptamers (Apts) are synthetic nucleic acid ligands that can be engineered to target various molecules, including amino acids, proteins, and pharmaceuticals. Through a series of adsorption, recovery, and amplification steps, Apts are extracted from combinatorial libraries of synthesized nucleic acids. Using aptasensors in bioanalysis and biomedicine can be improved by combining them with nanomaterials. Moreover, Apt-associated nanomaterials, including liposomes, polymeric, dendrimers, carbon nanomaterials, silica, nanorods, magnetic NPs, and quantum dots (QDs), have been widely used as promising nanotools in biomedicine. Following surface modifications and conjugation with appropriate functional groups, these nanomaterials can be successfully used in aptasensing. Advanced biological assays can use Apts immobilized on QD surfaces through physical interaction and chemical bonding. Accordingly, modern QD aptasensing platforms rely on interactions between QDs, Apts, and targets to detect them. QD-Apt conjugates can be used to directly detect prostate, ovarian, colorectal, and lung cancers or simultaneously detect biomarkers associated with these malignancies. Tenascin-C, mucin 1, prostate-specific antigen, prostate-specific membrane antigen, nucleolin, growth factors, and exosomes are among the cancer biomarkers that can be sensitively detected using such bioconjugates. Furthermore, Apt-conjugated QDs have shown great potential for controlling bacterial infections such as Bacillus thuringiensis, Pseudomonas aeruginosa, Escherichia coli, Acinetobacter baumannii, Campylobacter jejuni, Staphylococcus aureus, and Salmonella typhimurium. This comprehensive review discusses recent advancements in the design of QD-Apt bioconjugates and their applications in cancer and bacterial theranostics.