Functionalized-Graphene Field Effect Transistor-Based Biosensor for Ultrasensitive and Label-Free Detection of ß-Galactosidase Produced by Escherichia coli.

The detection of ß-galactosidase (ß-gal) activity produced by Escherichia coli (E. coli) can quickly analyze the pollution degree of seawater bodies in bathing and fishing grounds to avoid large-scale outbreaks of water pollution. Here, a functionalized biosensor based on graphene-based field effect transistor (GFET) modified with heat-denatured casein was developed for the ultrasensitive and label-free detection of the ß-gal produced by E. coli in real water samples. The heat-denatured casein coated on the graphene surface, as a probe linker and blocker, plays an important role in fabricating GEFT biosensor. The GFET biosensor response to the ß-gal produced by E. coli has a wide concentration dynamic range spanning nine orders of magnitude, in a concentration range of 1 fg·mL-1-100 ng·mL-1, with a limit of detection (LOD) 0.187 fg·mL-1 (1.61 aM). In addition to its attomole sensitivity, the GFET biosensor selectively recognized the ß-gal in the water sample and showed good selectivity. Importantly, the detection process of the ß-gal produced by E. coli can be completed by a straightforward one-step specific immune recognition reaction. These results demonstrated the usefulness of the approach, meeting environmental monitoring requirements for future use.

Methylation specific enzyme-linked oligonucleotide assays (MS-ELONA) for ultrasensitive DNA methylation analysis.

Methylation of the promoter region of cancer related genes plays a crucial role in the occurrence and development of cancer, and the degree of methylation has great potential for the early cancer diagnosis. At present, the technology used to quantify DNA methylation is mainly based on the DNA sequencing which are time-consuming and high-cost in the relating application. We have developed an ultrasensitive method of methylation specific enzyme-linked oligonucleotide assays (MS-ELONA) to detect and quantify the level of DNA methylation. We could detect as little as 2 pg of methylated DNA in the 100000-fold excess of unmethylated genes, and discriminate prostate cancer from benign prostatic hyperplasia (BPH) and control with serum samples. We also demonstrate the reversibility of DNA methylation modification by treatment with demethylation drugs. With 16-channel electrochemical work station, our research reveals a simple and inexpensive method to quantify the methylation level of specially appointed genes, and have the potential to be applied in the clinical research.

A novel biosensor based on a bio-barcode for the detection of Mycobacterium tuberculosis.

Tuberculosis (TB), the second (after COVID-19) deadliest infectious killer, is a chronic infectious disease caused by infection with Mycobacterium tuberculosis (M.T.), where early diagnosis and management are the key to containing the condition. Here, we report a novel biosensor for the detection of M.T. DNA based on magnetic separation, urease catalysis and silicon nanowire field effect transistor (SiNW FET) detection. M.T. DNA is sequence-specifically captured by magnetic nanoparticles and urease-labelled silica nanoparticles simultaneously to form a sandwich complex and urea is catalyzed into ammonium carbonate by urease modified on a sandwich complex. By using SiNW FET, the detection of M.T. DNA is realized indirectly by the detection of ammonium carbonate. The limit of detection (LOD) was determined to be 78.541 fM. The specificity of the biosensor was confirmed by detecting a panel of bacterial species. The utility of the biosensor was demonstrated in real-sample analysis and the recovery study of M.T. DNA was done in the genomic DNA extracted from cultured Mycobacterium tuberculosis. The biosensor holds promise to become a rapid, sensitive and accurate method for clinical diagnosis.

A DNA framework-based dual signal amplification biosensor for portable detection of SARS-CoV-2 and its mutations.

We developed a rapid and accurate biosensor to detect SARS-CoV-2 and distinguish its mutations. Benefitting from a DNA framework-modified ordered interface and a dual signal amplification strategy, our biosensor could detect SARS-CoV-2 with a detection limit down to 10 fM. It performed well on pseudo virus and SARS-CoV-2 RNA standard materials, revealing the potential application in disease diagnosis and spread, in combination with a home-made smartphone.

Correction: A smartphone-based three-in-one biosensor for co-detection of SARS-CoV-2 viral RNA, antigen and antibody.

Correction for 'A smartphone-based three-in-one biosensor for co-detection of SARS-CoV-2 viral RNA, antigen and antibody' by Yanzhi Dou et al., Chem. Commun., 2022, DOI: https://doi.org/10.1039/d2cc01297a.

A smartphone-based three-in-one biosensor for co-detection of SARS-CoV-2 viral RNA, antigen and antibody.

Rapid and comprehensive diagnostic methods are necessary for early identification and monitoring of SARS-CoV-2. Here, we have developed a universal and portable three-in-one biosensor linked to a smartphone for co-detection of SARS-CoV-2 viral RNA, antigen, and antibody. In combination with a smartphone, the online monitoring of SARS-CoV-2 virus-infected patients from infection to immunization could be intelligently achieved.

A Portable Biosensor Based on Au Nanoflower Interface Combined with Electrochemical Immunochromatography for POC Detection of Prostate-Specific Antigen.

Serum prostate-specific antigen (PSA) is a widely used for the detection of prostate cancer and is considered the most reliable biomarker. However, the currently reported detection methods cannot achieve rapid monitoring. Here, we report a novel electrochemical immunochromatography (EIC) system for clinically accurate PSA detection. First, we constructed a carbon interface modified with gold nanoflowers (Au NFs) based on screen-printed carbon electrodes (SPCE), which acted as nanostructures with larger specific surface area that increased the number of PSA capture antibodies and can further improve detection signal-to-noise (S/N) ratio. Then, we fabricated detection chips by combining the SPCE/Au NFs with EIC. Under optimized conditions, the proposed biosensor exhibits high accuracy, taking only 15 minutes to complete detection. By measuring the levels of PSA in clinical blood samples, the biosensor can successfully discriminate clinically diagnosed prostate cancer patients from healthy controls.

Smartphone-Based Electrochemical Biosensors for Directly Detecting Serum-Derived Exosomes and Monitoring Their Secretion.

Exosomes are potential biomarkers, which play an important role in early diagnosis and prognosis prediction of cancer-related diseases. Nevertheless, direct quantification of exosomes in biological fluid, especially in point-of-care tests (POCTs), remains extremely challenging. Herein, we developed a sensitive and portable electrochemical biosensor in combination with smartphones for quantitative analysis of exosomes. The improved double-antibody sandwich method-based poly-enzyme signal amplification was adopted to detect exosomes. We could detect as low as 7.23 ng of CD63-positive exosomes in 5 µL of serum within 2 h. Importantly, we demonstrated that the biosensor worked well with microliter-level serum and cell culture supernatant. The biosensor holds great potential for the detection of CD-63-expressing exosomes in early diagnosis of prostate disease because CD63-positive exosomes were less detected from the prostate patient serum. Also, the biosensor was used to monitor the secretion of exosomes with the drug therapy, showing a close relationship between the secretion of exosomes and the concentration of cisplatin. The biosensing platform provides a novel way toward POCT for the diagnosis and prognosis prediction of prostate disease and other diseases via biomarker expression levels of exosomes.

A nano-integrated microfluidic biochip for enzyme-based point-of-care detection of creatinine.

A nano-integrated portable enzymatic microfluidic electrochemical biochip was developed for single-step point-of-care testing of creatinine. The biochip could automatically eliminate a lot of interferences from practical biological samples and enzymatic intermediate products. Gold nanostructure- and carbon nanotube-based screen-printed carbon electrodes were integrated into microfluidic structures to improve the detection performance for creatinine. The microfluidic electrochemical biochip holds promise to become a practical device for medical diagnosis, especially POCT.

A Carbon-Based Antifouling Nano-Biosensing Interface for Label-Free POCT of HbA1c.

Electrochemical biosensing relies on electron transport on electrode surfaces. However, electrode inactivation and biofouling caused by a complex biological sample severely decrease the efficiency of electron transfer and the specificity of biosensing. Here, we designed a three-dimensional antifouling nano-biosensing interface to improve the efficiency of electron transfer by a layer of bovine serum albumin (BSA) and multi-walled carbon nanotubes (MWCNTs) cross-linked with glutaraldehyde (GA). The electrochemical properties of the BSA/MWCNTs/GA layer were investigated using both cyclic voltammetry and electrochemical impedance to demonstrate its high-efficiency antifouling nano-biosensing interface. The BSA/MWCNTs/GA layer kept 92% of the original signal in 1% BSA and 88% of that in unprocessed human serum after a 1-month exposure, respectively. Importantly, we functionalized the BSA/MWCNTs/GA layer with HbA1c antibody (anti-HbA1c) and 3-aminophenylboronic acid (APBA) for sensitive detection of glycated hemoglobin A (HbA1c). The label-free direct electrocatalytic oxidation of HbA1c was investigated by cyclic voltammetry (CV). The linear dynamic range of 2 to 15% of blood glycated hemoglobin A (HbA1c) in non-glycated hemoglobin (HbAo) was determined. The detection limit was 0.4%. This high degree of differentiation would facilitate a label-free POCT detection of HbA1c.

A Carbon-Based DNA Framework Nano-Bio Interface for Biosensing with High Sensitivity and a High Signal-to-Noise Ratio.

Biosensing interface based on screen-printed carbon electrodes (SPCE) has been widely used for electrochemical biosensors in the field of medical diagnostics, food safety, and environmental monitoring. Nevertheless, SPCE always has a rough surface, which is easy to result in the disorder of nucleic acid capture probes, the nonspecific adsorption of signaling probes, the steric hindrance of target binding, and decrease in the signal-to-noise ratio and sensitivity of biosensors. So far, it still remains extremely challenging to develop high-efficiency carbon-based biosensing interfaces, especially for DNA probe-based assembly and functionalization. In this paper, we first used a specific DNA framework, DNA tetrahedron to solve the defects of the carbon interface, improving the biosensing ability of SPCE. With covalent coupling, the DNA tetrahedron could be immobilized on the carbon surface. Biosensing probe sequences extending from the DNA tetrahedron can be changed for different target molecules. We demonstrated that the improved SPCE could be applied for the detection of a variety of bioactive molecules. Typically, we designed gap hybridization, aptamer "sandwich" and aptamer competition reduction strategy for the detection of miRNA-141, thrombin, and ATP, respectively. High signal-to-noise ratio, sensitivity, and specificity were obtained for all of these kinds. Especially, the DNA tetrahedron-modified SPCE can work well with serum samples. The carbon-based DNA framework nano-bio interface would expand the use of SPCE and make electrochemical biosensors more available and valuable in clinical diagnosis.

DNA Framework-Supported Electrochemical Analysis of DNA Methylation for Prostate Cancers.

Epigenetic alterations hold great promise as biomarkers for early stage cancer diagnosis. Nevertheless, direct identification of rare methylated DNA in the genome remains challenging. Here, we report an ultrasensitive framework nucleic acid-based electrochemical sensor for quantitative and highly selective analysis of DNA methylation. Notably, we can detect 160 fg of methylated DNA in million-fold unmethylated DNA samples using this electrochemical methylation-specific polymerase chain reaction (E-MSP) method. The high sensitivity of E-MSP enables one-step detection of low-abundance methylation at two different genes in patient serum samples. By using a combination test with two methylation alterations, we achieve high accuracy and sensitivity for reliable differentiation of prostate cancer and benign prostate hypertrophy (BPH). This new method sheds new light on translational use in early cancer diagnosis and in monitoring patients' responses to therapeutic agents.

Multicolor Gold-Silver Nano-Mushrooms as Ready-to-Use SERS Probes for Ultrasensitive and Multiplex DNA/miRNA Detection.

Uniform silver-containing metal nanostructures with strong and stable surface-enhanced Raman scattering (SERS) signals hold great promise for developing ultrasensitive probes for biodetection. Nevertheless, the direct synthesis of such ready-to-use nanoprobes remains extremely challenging. Herein we report a DNA-mediated gold-silver nanomushroom with interior nanogaps directly synthesized and used for multiplex and simultaneous SERS detection of various DNA and RNA targets. The DNA involved in the nanostructures can act as not only gap DNA (mediated DNA) but also probe DNA (hybridized DNA), and DNA's involvement enables the nanostructures to have the inherent ability to recognize DNA and RNA targets. Importantly, we were the first to establish a new method for the generation of multicolor SERS probes using two different strategies. First Raman-labeled alkanethiol probe DNA was assembled on gold nanoparticles, and second, thiol-containing Raman reporters were coassembled with the probe DNA. The ready-to-use probes also give great potential to develop ultrasensitive detection methods for various biological molecules.

High-Sensitivity and High-Efficiency Detection of DNA Hydroxymethylation in Genomic DNA by Multiplexing Electrochemical Biosensing.

DNA hydroxymethylation (5-hmC) is a kind of new epigenetic modification, which plays key roles in DNA demethylation, genomic reprogramming, and the gene expression in mammals. For further exploring the functions of 5-hmC, it is necessary to develop sensitive and selective methods for detecting 5-hmC. Herein, we developed a novel multiplexing electrochemical (MEC) biosensor for 5-hmC detection based on the glycosylation modification of 5-hmC and enzymatic signal amplification. The 5-hmC was first glycosylated by T4 ß-glucosyltransferase and then oxidated by sodium periodate. The resulting glucosyl-modified 5-hmC (5-ghmC) was incubated with ARP-biotin and was bound to avidin-HRP. The 5-hmC can be detected at the subnanogram level. Finally, we performed 5-hmC detection for mouse tissue samples and cancer cell lines. The limit of detection of the MEC biosensor is 20 times lower than that of commercial kits based on optical meaurement. Also, the biosensor presented high detection specificity because the chemical reaction for 5-hmC modification can not happen at any other unhydroxymethylated nucleic acid bases. Importantly, benefited by its multiplexing capacity, the developed MEC biosensor showed excellent high efficiency, which was time-saving and cost less.

Unusual Fe(CN)6³â»/4⁻ capture induced by synergic effect of electropolymeric cationic surfactant and graphene: characterization and biosensing application.

Herein, a special microheterogeneous system for Fe(CN)6(3-/4-) capture was constructed based on graphene (GN) and the electropolymeric cationic surfactant, an amphiphilic pyrrole derivative, (11-pyrrolyl-1-yl-undecyl) triethylammonium tetrafluoroborate (A2). The morphology of the system was characterized by scanning electron microscope. The redox properties of the entrapped Fe(CN)6(3-/4-) were investigated by cyclic voltammetry and UV-visible spectrometry. The entrapped Fe(CN)6(3-/4-) exhibited highly electroactive with stable and symmetrical cyclic voltammetric signal. A dramatic negative shift in the half wave potential can be obtained due to the unusual Fe(CN)6(3-/4-) partitioning in in this microheterogeneous system based on poly(A2+GN). Finally, the entrapped Fe(CN)6(3-/4-) was applied in the construction of the enhanced biosensors to hydrogen peroxide and sulfide.