Disposable electrocatalytic sensor for whole blood NADH monitoring.

Monitoring nicotinamide adenine dinucleotide (NADH) is important because NADH is involved in cellular redox reactions and cellular energy production. Currently, few biosensors quantify NADH in whole blood. However, they still have limitations due to several defects, including poor repeatability, long analysis time, and their requirement of extra sample pretreatment. In this study, we developed electrocatalytic sensors using screen-printed electrodes with a redox-active monolayer 4'-mercapto-N-phenylquinone diamine formed by a self-assembled monolayer of a 4-aminothiophenol (4-ATP). We exhibited their behavior as electrocatalysts toward the oxidation of NADH in whole blood. Finally, the electrocatalytic sensors maintained stability and exhibited 3.5 µM limit of detection, with 0.0076 ± 0.0006 µM/µA sensitivity in a mouse's whole blood. As proof of concept, a polyhexamethylene guanidine phosphate-treated mouse model was used to induce inflammatory and fibrotic responses, and NADH level was measured for 45 days. This work demonstrates the potential of electrocatalytic sensors to analyze NADH in whole blood and to be developed for extensive applications.

An immunosensor based on a high performance dual-gate oxide semiconductor thin-film transistor for rapid detection of SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the agent of an infectious disease that has led the WHO to declare its highest level (6) pandemic. The coronavirus disease 2019 (COVID-19) has spread rapidly around the world, and the number of confirmed cases has passed 246 million as of November 2021. Therefore, precise and fast virus detection protocols need to be developed to cope with the rapid spread of the virus. Here, we present a high performance dual-gate oxide semiconductor thin-film transistor (TFT)-based immunosensor for detecting SARS-CoV-2. The immunosensor has an indium tin oxide sensing membrane to which the antibody against the SARS-CoV-2 spike S1 protein can be immobilized through functionalization. The dual-gate TFT was stable under ambient conditions with near-zero hysteresis; capacitive coupling yields a 10.14 ± 0.14-fold amplification of the surface charge potential on the sensing membrane and improves the pH sensitivity to 770.1 ± 37.74 mV pH-1 above the Nernst limit. The immunosensor could rapidly detect the SARS-CoV-2 spike S1 protein and cultured SARS-CoV-2 in 0.01× PBS with high antigen selectivity and sensitivity. Our immunosensor can accurately measure the electrical changes originated from SARS-CoV-2, without the need for polymerase chain reaction tests or labeling.

Carbon nanotube field-effect transistor (CNT-FET)-based biosensor for rapid detection of SARS-CoV-2 (COVID-19) surface spike protein S1.

The large-scale diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is important for traceability and treatment during pandemic outbreaks. We developed a fast (2-3 min), easy-to-use, low-cost, and quantitative electrochemical biosensor based on carbon nanotube field-effect transistor (CNT-FET) that allows digital detection of the SARS-CoV-2 S1 in fortifited saliva samples for quick and accurate detection of SARS-CoV-2 S1 antigens. The biosensor was developed on a Si/SiO2 surface by CNT printing with the immobilization of a anti-SARS-CoV-2 S1. SARS-CoV-2 S1 antibody was immobilized on the CNT surface between the S-D channel area using a linker 1-pyrenebutanoic acid succinimidyl ester (PBASE) through non-covalent interaction. A commercial SARS-CoV-2 S1 antigen was used to characterize the electrical output of the CNT-FET biosensor. The SARS-CoV-2 S1 antigen in the 10 mM AA buffer pH 6.0 was effectively detected by the CNT-FET biosensor at concentrations from 0.1 fg/mL to 5.0 pg/mL. The limit of detection (LOD) of the developed CNT-FET biosensor was 4.12 fg/mL. The selectivity test was performed by using target SARS-CoV-2 S1 and non-target SARS-CoV-1 S1 and MERS-CoV S1 antigens in the 10 mM AA buffer pH 6.0. The biosensor showed high selectivity (no response to SARS-CoV-1 S1 or MERS-CoV S1 antigen) with SARS-CoV-2 S1 antigen detection in the 10 mM AA buffer pH 6.0. The biosensor is highly sensitive, saves time, and could be a helpful platform for rapid detection of SARS-CoV-2 S1 antigen from the patients saliva.

Penta-peptide ATN-161 based neutralization mechanism of SARS-CoV-2 spike protein.

SARS-CoV-2 has become a big challenge for the scientific community worldwide. SARS-CoV-2 enters into the host cell by the spike protein binding with an ACE2 receptor present on the host cell. Developing safe and effective inhibitor appears an urgent need to interrupt the binding of SARS-CoV-2 spike protein with ACE2 receptor in order to reduce the SARS-CoV-2 infection. We have examined the penta-peptide ATN-161 as potential inhibitor of ACE2 and SARS-CoV-2 spike protein binding, where ATN-161 has been commercially approved for the safety and possess high affinity and specificity towards the receptor binding domain (RBD) of S1 subunit in SARS-CoV-2 spike protein. We carried out experiments and confirmed these phenomena that the virus bindings were indeed minimized. ATN-161 peptide can be used as an inhibitor of protein-protein interaction (PPI) stands as a crucial interaction in biological systems. The molecular docking finding suggests that the binding energy of the ACE2-spike protein complex is reduced in the presence of ATN-161. Protein-protein docking binding energy (-40.50 kcal/mol) of the spike glycoprotein toward the human ACE2 and binding of ATN-161 at their binding interface reduced the biding energy (-26.25 kcal/mol). The finding of this study suggests that ATN-161 peptide can mask the RBD of the spike protein and be considered as a neutralizing candidate by binding with the ACE2 receptor. Peptide-based masking of spike S1 protein (RBD) and its neutralization is a highly promising strategy to prevent virus penetration into the host cell. Thus masking of the RBD leads to the loss of receptor recognition property which can reduce the chance of infection host cells.

Review: Roles of human serum albumin in prediction, diagnoses and treatment of COVID-19.

The severe acute respiratory syndrome corona virus-2 (SARS-CoV-2) keeps on destroying normal social integrity worldwide, bringing about extraordinary medical services, cultural and financial interruption. Individuals with diabetes have been demonstrated to be at higher risk of complications and even death when exposed to SARS-CoV-2. Regardless of pandemic scale infection, there is presently limited comprehension on the potential impact of SARS-CoV-2 on individuals with diabetes. Human serum albumin (HSA) is the most abundant circulating plasma protein in human serum and attracted more interest from researchers because most susceptible to non-enzymatic glycation reactions. Albumin down-regulates the expression of ACE2 that is the target receptor of COVID-19. Hypoalbuminemia, coagulopathy, and vascular disease have been connected in COVID-19 and appear to predict outcomes independent of age and morbidity. This review discusses the most recent evidence that the ACE/ACE2 ratio could influence by human serum albumin both the susceptibility of individuals to SARS-CoV-2 infection and the outcome of the COVID-19 disease.

Structure, enzymatic activities, glycation and therapeutic potential of human serum albumin: A natural cargo.

Human serum albumin (HSA) is an opulent, non-glycosylated, most versatile carrier protein in plasma possessing multiple functions. HSA has the ability to interact with a variety of ligands, including exogenous pharmacological drugs. HSA has multiple binding sites located in different subdomains and which are responsible for binding of ligands. While antecedent research has discovered various functional and structural properties of HSA, the objective of this review paper is to shed light on some of the important properties of HSA and how binding pattern of different ligands can sustain the development of new drugs. Some significant properties include transportation, ligand-binding, distribution and metabolism of a compound. The HSA molecule can undergo various structural changes modifying its conformation and finally affects the ligand binding properties and redox state. Another important feature is an esterase-like activity possessed by HSA, which is also crucial in converting the prodrugs into active therapeutics. Therefore, HSA is one of the most suitable molecules for future research in drug discovery in pharmaceutical industry because of its numerous features and binding pattern that also governs the metabolism and drug dosage.

Purification of semiconducting single-walled carbon nanotubes by spiral counter-current chromatography.

Over the last decade man-made carbon nanostructures have shown great promise in electronic applications, but they are produced as very heterogeneous mixtures with different properties so the achievement of a significant commercial application has been elusive. The dimensions of single-wall carbon nanotubes are generally a nanometer wide, up to hundreds of microns long and the carbon nanotubes have anisotropic structures. They are processed to have shorter lengths but they need to be sorted by diameter and chirality. Thus counter-current chromatography methods developed for large molecules are applied to separate these compounds. A modified mixer-settler spiral CCC rotor made with 3 D printed disks was used with a polyethylene glycol-dextran 2-phase solvent system and a surfactant gradient to purify the major species in a commercial preparation. We isolated the semi-conducting single walled carbon nanotube chiral species identified by UV spectral analysis. The further development of spiral counter-current chromatography instrumentation and methods will enable the scalable purification of carbon nanotubes useful for the next generation electronics.

Spiral Countercurrent Chromatography Enrichment, Characterization, and Assays of Carbon Nanotube Chiralities for Use in Biosensors.

Single-walled carbon nanotubes (SWCNTs) are synthetic materials that hold great promise for electronics that are smaller and more versatile than the current silica-based technologies. But as-produced SWCNTs are generally a mixture of nanotubes with different structures that have vastly different properties. Separating these SWCNTs from multiwalled and metallic carbon nanotubes is vital to explore their individual properties and commercial utility ranging from optics to semiconductors. Compounding the problem of commercial investigation is that the semiconducting SWCNTs are also a mixture of different diameters and/or chiralities with different properties. Analyzing properties of enriched semiconducting SWCNT chiralities has only recently been possible through separation techniques such as aqueous two-phase solvent systems. Our study illustrates a semipreparative spiral countercurrent chromatography (CCC) separation of a commercial mixture of SWCNTs into distinct enriched fractions. A new mixer-settler spiral disk rotor was applied in this study, in which we compare the enriched SWCNTs for their effectiveness in biosensors with a high-throughput model assay, followed by antibody-mediated detection of Escherichia coli. Our results demonstrate that CCC-enriched responsive SWCNTs for biosensors can be used in our model assay, as well as for the detection of E. coli. To date, we believe that this is the first study along with Liu et al. [Chirality-controlled synthesis of single-wall carbon nanotubes using vapour-phase epitaxy. Nat. Commun.2012, 3, 1199] to demonstrate a specific utility of separated SWCNT species.

High-throughput Peptide epitope mapping using carbon nanotube field-effect transistors.

Label-free and real-time detection technologies can dramatically reduce the time and cost of pharmaceutical testing and development. However, to reach their full promise, these technologies need to be adaptable to high-throughput automation. To demonstrate the potential of single-walled carbon nanotube field-effect transistors (SWCNT-FETs) for high-throughput peptide-based assays, we have designed circuits arranged in an 8 × 12 (96-well) format that are accessible to standard multichannel pipettors. We performed epitope mapping of two HIV-1 gp160 antibodies using an overlapping gp160 15-mer peptide library coated onto nonfunctionalized SWCNTs. The 15-mer peptides did not require a linker to adhere to the non-functionalized SWCNTs, and binding data was obtained in real time for all 96 circuits. Despite some sequence differences in the HIV strains used to generate these antibodies and the overlapping peptide library, respectively, our results using these antibodies are in good agreement with known data, indicating that peptides immobilized onto SWCNT are accessible and that linear epitope mapping can be performed in minutes using SWCNT-FET.

Comparison of radioimmuno and carbon nanotube field-effect transistor assays for measuring insulin-like growth factor-1 in a preclinical model of human breast cancer.

BACKGROUND: To realize the promise of personalized medicine, diagnostic instruments used for detecting and measuring biomarkers must become smaller, faster and less expensive. Although most techniques used currently to detect biomarkers are sensitive and specific, many suffer from several disadvantages including their complexity, high cost and long turnaround time. One strategy to overcome these problems is to exploit carbon nanotube (CNT) based biosensors, which are sensitive, use inexpensive disposable components and can be easily adapted to current assay protocols. In this study we investigated the applicability of using a CNT field-effect transistor (CNT-FET) as a diagnostic instrument for measuring cancer biomarkers in serum using a mouse model of Breast Cancer Susceptibility 1-related breast cancer. Insulin like growth factor-1 (IGF-1) was chosen because it is highly relevant in breast cancer and because measuring serum IGF-1 levels by conventional methods is complicated due to specific IGF-1 serum binding proteins. FINDINGS: Our results show that there is good correlation between the two platforms with respect to detecting serum IGF-1. In fact, the CNT-FETs required only one antibody, gave real-time results and required approximately 100-fold less mouse serum than the radioimmunoassay. CONCLUSIONS: Both IGF-1 radioimmuno and CNT-FET assays gave comparable results. Indeed, the CNT-FET assay was simpler and faster than the radioimmunoassay. Additionally, the low serum sample required by CNT-FETs can be especially advantageous for studies constricted by limited amount of human clinical samples and for mouse studies, since animals often need to be sacrificed to obtain enough serum for biomarker evaluation.

Evidence for an additional metastatic route: in vivo imaging of cancer cells in the primo-vascular system around tumors and organs.

PURPOSE: Researchers have been studying the mechanisms by which metastasis can be prevented via blocking the hematogenous and the lymphatic routes for a long time now. However, metastasis is still the single most challenging obstacle for successful cancer management. In a new twist that may require some retooling of this established approach, we investigated the hypothesis that tumor metastases can occur via an independent fluid-conducting system called the primo-vascular system. PROCEDURES: The dissemination and growth of near-infrared quantum dot (NIR QD)-electroporated cancer cells in metastatic sites were investigated using in vivo multispectral imaging techniques. RESULTS: Our results show that the NIR QD-labeled cancer cells were able to migrate through not only the blood vascular and lymphatic systems but also the primo-vascular system extending from around the tumor to inside the abdominal cavity. Furthermore, the NIR QD-labeled cancer cells, which had been seeded intraperitoneally, specifically infiltrated the primo-vascular system in the omentum and in the gonadal fat. CONCLUSIONS: These findings strongly suggest that the primo-vascular system may be an additional metastasis route, complementing the lymphatic and hematogenous routes, which facilitate the dissemination and colonization of cancer cells at secondary sites.

Bonghan ducts as possible pathways for cancer metastasis.

OBJECTIVE: The present study has been designed to find a possible new route for the metastasis of cancer cells on the fascia surrounding tumor tissue using a novel technique of trypan blue staining. MATERIALS AND METHODS: Tumor tissues were grown in the skin of nude mice after sub-cutaneous inoculation with human lung cancer cells. Trypan blue was recently identified as a dye with specificity for Bonghan ducts (BHDs) and not other tissues, such as blood or lymph vessels or nerves. RESULTS: We demonstrate that the trypan blue staining technique allows the first visualization of BHDs which are connected to tumor tissues. CONCLUSION: Since BHDs are known to make up a circulatory system corresponding to acupuncture meridians or collaterals, we propose that, in addition to the currently known blood or lymph vessels, BHDs on tumor tissue fascia may be a novel pathway for metastasis.

Enhancement of ultraweak photon emission with 3 MHz ultrasonic irradiation on transplanted tumor tissues of mice.

We investigated photon emissions of various bio-samples which were induced by ultrasonic stimulation. It has been reported that ultrasonic stimulations induced the thermal excitation of the bio-tissues. After ultrasonic stimulation, any measurement of photon radiation in the visible spectral range has not been carried out yet. The instruments consisted of electronic devices for an ultrasonic generator of the frequency 3 MHz and a photomultiplier tube (PMT) system counting photons from bio-tissues. The transplanted tumor tissues of mice were prepared for the experiments and their liver and spleen tissues were also used for the controls. It was found that the continuous ultrasonic stimulations with the electrical power 2300 mW induced ultraweak photon emissions from the tumor tissues. The number of induced photon was dependent of the type of the tissues and the stimulation time intervals. The level of photon emission was increased from the mouse tumor exposed to the ultrasonic stimulations, and the changes were discriminated from those of the spleens and livers.

Evaluation of corrosion behaviors and surface profiles of platinum-coated electrodes by electrochemistry and complementary microscopy: biomedical implications for anticancer therapy.

Corrosion behaviors and surface profiles of platinum (Pt)-coated electrodes were investigated by electrochemistry and complementary microscopy. Four types of needle electrodes (Pt/titanium, Pt/tungsten, Pt/brass, and Pt/stainless steel) were prepared by unbalanced magnetron sputtering and subjected to the potentiodynamic polarization test in 0.9% deaerated sodium chloride solution at room temperature. The potentiodynamic test revealed that Pt/Ti electrodes had the highest corrosion resistance among the electrodes tested in this study, with the corrosion potential of 228mV(SCE) and the corrosion rate of 0.10 microA/cm(2). Morphology and surface profiles of the electrodes examined by reflected light, electron, and atomic force microscopy were consistent with those of corrosion resistance, showing that the electrodes with higher corrosion resistance had lower surface roughness than others. It is likely that the electrodes with higher corrosion resistance have suffered slower dissolution of substrates, and consequently had less cavities and/or corrosion products. Furthermore, nude mice inoculated with a bronchoalveolar cancer cell line and exposed to direct electric field showed the deterioration of proliferated tumor cells, proving the efficacy of electrochemical treatment. These results suggest that either Pt/Ti or Pt/W electrodes for solid tumors are suitable to be employed in the electrochemical treatment of tumors for anticancer therapy, applying low-level direct current due to their high corrosion resistance.

Scanning spontaneous photon emission from transplanted ovarian tumor of mice using a photomultiplier tube.

A scanning system for the detection of spontaneous ultraweak photon emission from nude mice with transplanted tumors is presented. A photomultiplier tube (PMT) with an effective area of 15 mm diameter was used for measuring photon emission in a wavelength range from 300 to 650 nm. Tumors were induced in nude mice by transplantation of an ovarian cancer cell line into the back of mice. The PMT was moved for scanning over the whole body of a mouse placed in a dark box. The profiles of the intensities of photon emissions from the tumor mice are presented and compared with those obtained from the control mice.

Label free novel electrical detection using micromachined PZT monolithic thin film cantilever for the detection of C-reactive protein.

In this paper, we report the novel electrical measurement for the label-free detection of C-reactive protein (CRP) using resonant frequency shift in the monolithic thin film cantilever of micromachined Pb(Zr0.52Ti0.48)O3 (PZT) which was fabricated with the composition of SiO2/Ta/Pt/PZT/Pt/SiO2 on silicon nitride (SiNx) supporting layer for the dual purpose of electrical self-excitation and sensing. The specific binding characteristics of CRP antigen to its antibody, which is immobilized with Calixcrown SAMs on Au surface deposited on microcantilever, is determined in high sensitivity to the nanogram level per milliliter by measuring the resonant frequency shift. The nanomechanical PZT cantilever turns out a robust platform for the highly specific antigen-antibody interaction and provides with the novel tool for qualification and quantification of biomolecules without any sample labeling and bulky optical apparatus.