Detecting exosomes specifically: a multiplexed device based on alternating current electrohydrodynamic induced nanoshearing

Exosomes show promise as non-invasive biomarkers for cancers, but their effective capture and specific detection is a significant challenge. Herein, we report a multiplexed microfluidic device for highly specific capture and detection of multiple exosome targets using a tuneable alternating current electrohydrodynamic (ac-EHD) methodology - referred to as nanoshearing. In our system, electrical body forces generated by ac-EHD act within nanometers of an electrode surface (i.e., within the electrical layer) to generate nanoscaled fluid flow which enhances the specificity of capture and also reduce nonspecific adsorption of weakly bound molecules from the electrode surface. This approach demonstrates the analysis of exosomes derived from cells expressing human epidermal growth factor receptor 2 (HER2) and prostate specific antigen (PSA), and exhibits a 5-fold detection enhancement compared to hydrodynamic flow based assays. The device was also sensitive enough to detect approximately 2750 exosomes/µL (n = 3) and also capable of specifically isolating exosomes from breast cancer patient samples. We believe this approach can potentially find its relevance as a simple and rapid quantification tool to analyze exosome targets in biological applications

Electrochemical detection of glycan and protein epitopes of glycoproteins in serum

Aberrant protein glycosylation is associated with a range of pathological conditions including cancer and possesses diagnostic importance. Translation of glycoprotein biomarkers will be facilitated by the development of a rapid and sensitive analytical platform that simultaneously interrogates both the glycan and protein epitopes of glycoproteins in body fluids such as serum or saliva. To this end, we developed an electrochemical biosensor based on the immobilization of a lectin on the gold electrode surface to recognize/capture a target glycan epitope conjugated to glycoproteins, followed by detection of the protein epitope using a target protein-specific antibody. Electrochemical signals are generated by label-free voltammetric or impedimetric interrogation of a ferro/ferricyanide redox couple (e.g. [Fe(CN)(6)](3-/4-)) on the sensing surface, where the change in voltammetric current or interfacial electron transfer resistance was measured. The detection system was demonstrated using the model glycoprotein chicken ovalbumin with Sambucus nigra agglutinin type I (SNA lectin), and exhibits femtomolar sensitivity in the background of diluted human serum. The results obtained in this proof-of-concept study demonstrate the possibility of using electrochemical detection for developing cheap point-of-care diagnostics with high specificity and sensitivity for blood glycoprotein biomarkers

Sustainable Antibiotic-Free Broiler Meat Production: Current Trends, Challenges, and Possibilities in a Developing Country Perspective

Antibiotic-free broiler meat production is becoming increasingly popular worldwide due to consumer perception that it is superior to conventional broiler meat. Globally, broiler farming impacts the income generation of low-income households, helping to alleviate poverty and secure food in the countryside and in semi-municipal societies. For decades, antibiotics have been utilized in the poultry industry to prevent and treat diseases and promote growth. This practice contributes to the development of drug-resistant bacteria in livestock, including poultry, and humans through the food chain, posing a global public health threat. Additionally, consumer demand for antibiotic-free broiler meat is increasing. However, there are many challenges that need to be overcome by adopting suitable strategies to produce antibiotic-free broiler meat with regards to food safety and chicken welfare issues. Herein, we focus on the importance and current scenario of antibiotic use, prospects, and challenges in the production of sustainable antibiotic-free broiler meat, emphasizing broiler farming in the context of Bangladesh. Moreover, we also discuss the need for and challenges of antibiotic alternatives and provide a future outlook for antibiotic-free broiler meat production

Detection of regional DNA methylation using DNA-graphene affinity interactions

We report a new multiplexed strategy for the electrochemical detection of regional DNA methylation across multiple regions. Using the sequence dependent affinity of bisulfite treated DNA towards gold surfaces, the method integrates the high sensitivity of a micro-fabricated multiplex device comprising a microarray of gold electrodes, with the powerful multiplexing capability of multiplex-PCR. The synergy of this combination enables the monitoring of the methylation changes across several genomic regions simultaneously from as low as 500 pg μl(-1) of DNA with no sequencing requirement

eMethylsorb: rapid quantification of DNA methylation in cancer cells on screen-printed gold electrodes

Simple, sensitive and inexpensive regional DNA methylation detection methodologies are imperative for routine patient diagnostics. Herein, we describe eMethylsorb, an electrochemical assay for quantitative detection of regional DNA methylation on a single-use and cost-effective screen-printed gold electrode (SPE-Au) platform. The eMethylsorb approach is based on the inherent differential adsorption affinity of DNA bases to gold (i.e. adenine > cytosine ≥ guanine > thymine). Through bisulfite modification and asymmetric PCR of DNA, methylated and unmethylated DNA in the sample becomes guanine-enriched and adenine-enriched respectively. Under optimized conditions, adenine-enriched unmethylated DNA (higher affinity to gold) adsorbs more onto the SPE-Au surface than methylated DNA. Higher DNA adsorption causes stronger coulombic repulsion and hinders reduction of ferricyanide [Fe(CN)]ions on the SPE-Au surface to give a lower electrochemical response. Hence, the response level is directly proportional to the methylation level in the sample. The applicability of this methodology was tested by detecting the regional methylation status in a cluster of eight CpG sites within the engrailed (EN1) gene promoter of the MCF7 breast cancer cell line. A 10% methylation level sensitivity with good reproducibility (RSD = 5.8%, n = 3) was achieved rapidly in 10 min. Furthermore, eMethylsorb also has advantages over current methylation assays such as being inexpensive, rapid and does not require any electrode surface modification. We thus believe that the eMethylsorb assay could potentially be a rapid and accurate diagnostic assay for point-of-care DNA methylation analysis

Superparamagnetic nanoarchitectures for disease-specific biomarker detection

© 2019 The Royal Society of Chemistry. The detection of clinically relevant disease-specific biomolecules, including nucleic acids, circulating tumor cells, proteins, antibodies, and extracellular vesicles, has been indispensable to understand their functions in disease diagnosis and prognosis. Therefore, a biosensor for the robust, ultrasensitive, and selective detection of these low-Abundant biomolecules in body fluids (blood, urine, and saliva) is emerging in current clinical research. In recent years, nanomaterials, especially superparamagnetic nanomaterials, have played essential roles in biosensing due to their intrinsic magnetic, electrochemical, and optical properties. However, engineered multicomponent magnetic nanoparticle-based current biosensors that offer the advantages of excellent stability in a complex biomatrix; easy and alterable biorecognition of ligands, antibodies, and receptor molecules; and unified point-of-care integration have yet to be achieved. This review introduces the recent advances in superparamagnetic nanostructures for electrochemical and optical biosensing for disease-specific biomarkers. This review emphasizes the synthesis, biofunctionalization, and intrinsic properties of nanomaterials essential for robust, ultrasensitive biosensing. With a particular emphasis on nanostructure-based electrochemical and optical detection of disease-specific biomarkers such as nucleic acids (DNA and RNA), proteins, autoantibodies, and cells, this review also chronicles the needs and challenges of nanoarchitecture-based detection. These summaries provide further insights for researchers to inspire their future work on the development of nanostructures for integrating into biosensing and devices for a broad field of applications in analytical sensing and in clinic

Trace analysis of DNA: Preconcentration, separation, and electrochemical detection in microchip electrophoresis using Au nanoparticles

We have developed a simple and sensitive on-chip preconcentration, separation, and electrochemical detection (ED) method for trace analysis of DNA. The microchip comprised of three parallel channels: the first two are for the field-amplified sample stacking and subsequent field-amplified sampled injection steps, while the third one is for the microchip gel electrophoresis (MGE) with ED (MGE-ED). To improve preconcentration and separation performances of the method, the stacking and separation buffers containing the hydroxypropyl cellulose (HPC) matrix were modified with gold nanoparticles (AuNPs). The formation of AuNPs and HPC/AuNP-modified buffers were characterized by UV−visible spectroscopy and TEM experiments. The conducting polymer-modified electrode was also modified with AuNPs to enhance detection performances of the electrode. The conducting polymer/AuNP layers act as electrocatalysts for the direct detection of DNA based on their oxidation in a solution phase. The total sensitivity was improved by 25000-fold when compared with a conventional MGE-ED analysis. The calibration plots were linear (r2 = 0.9993) within the range of 0.003−1.0 pg/μL for a 20-bp DNA sample. The sensitivity was 0.20 nA/(fg/μL), with a detection limit of 5.7 amol in a 50-μL sample, based on S/N = 3. The applicability of the method for the analysis of 13 fragments present in a 100-bp DNA ladder was successfully demonstrated

Microchip and capillary electrophoresis using nanoparticles

Explores the latest applications arising from the intersection of nanotechnology and microfluidics In the past two decades, microfluidics research has seen phenomenal growth, with many new and emerging applications in fields ranging from chemistry, physics, and biology to engineering. With the emergence of nanotechnology, microfluidics is currently undergoing dramatic changes, embracing the rising field of nanofluidics. This volume reviews the latest devices and applications stemming from the merging of nanotechnology with microfludics in such areas as drug discovery, bio-sensing, catalysis, electrophoresis, enzymatic reactions, and nanomaterial synthesis. Each of the ten chapters is written by a leading pioneer at the intersection of nanotechnology and microfluidics. Readers not only learn about new applications, but also discover which futuristic devices and applications are likely to be developed. Topics explored in this volume include: * New lab-on-a-chip systems for drug delivery * Integration of microfluidics with nanoneuroscience to study the nervous system at the single-cell level * Recent applications of nanoparticles within microfluidic channels for electrochemical and optical affinity biosensing * Novel microfluidic approaches for the synthesis of nanomaterials * Next-generation alternative energy portable power devices References in each chapter guide readers to the primary literature for further investigation of individual topics. Overall, scientists, researchers, engineers, and students will not only gain a new perspective on what has been done, but also the nanotechnology tools they need to develop the next generation of microfluidic devices and applications. Microfluidic Devices for Nanotechnology is a two-volume publication, the first ever to explore the synergies between microfluidics and nanotechnology. The first volume covers fundamental concepts; this second volume examines applications

"Drill and fill" lithography for controlled fabrication of 3D platinum electrodes

Reproducible fabrication of three-dimensional (3D) metal electrodes is essential for accurate measurements of analyte in the fields of electroanalytical chemistry and biosensor development. Unfortunately, fabrication of these types of electrodes is complicated by the inherent complexity of current nano-fabrication methodologies. Herein, we report a simple and rapid fabrication method referred to as "drill and fill" lithography that provides control structuring and excellent reproducibility in the fabrication of 3D Pt electrodes with different sizes and shapes. The ability to combine "drill" and "fill" in a single technique using focused ion beam (FIB) lithography enabled precise control of the size and shape of the electrode depending on the amount of deposited metallic Pt. Electrodes were characterized using scanning electron microscopy, cyclic voltammetry, and Faradaic electrochemical impedance spectroscopy. The key innovation of this methodology lies in its simplicity and ability to tune the operating parameters to obtain electrodes of designated size and shapes. We envisage that these electrodes could be ideal for 'on-chip' diagnostic platforms