MicroRNAs (miRNAs) play several crucial roles in the physiological and pathological processes of the human body. They are considered as important biomarkers for the diagnosis of various disorders. Thus, rapid, sensitive, selective, and affordable detection of miRNAs is of great importance. However, the small size, low abundance, and highly similar sequences of miRNAs impose major challenges to their accurate detection in biological samples. In recent years, metal-organic frameworks (MOFs) have been applied as promising sensing materials for the fabrication of different biosensors due to their distinctive characteristics, such as high porosity and surface area, tunable pores, outstanding adsorption affinities, and ease of functionalization. In this review, the applications of MOFs and MOF-derived materials in the fabrication of fluorescence, electrochemical, chemiluminescence, electrochemiluminescent, and photoelectrochemical biosensors for the detection of miRNAs and their detection principle and analytical performance are discussed. This paper attempts to provide readers with a comprehensive knowledge of the fabrication and sensing mechanisms of miRNA detection platforms.
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.
Cervical cancer emerges as the third most prevalent types of malignancy among women on a global scale. Cervical cancer is significantly associated with the persistent infection of human papillomavirus (HPV) type 16. The process of diagnosing is crucial in order to prevent the progression of a condition into a malignant state. The early detection of cervical cancer through initial stage screening is of the utmost significance in both the prevention and effective management of this disease. The present detection methodology is dependent on quantitative polymerase chain reaction (qPCR), which necessitates the use of a costly heat cycler instrument. In this study, we report the development of an electrochemical DNA biosensor integrated with an isothermal recombinase polymerase amplification (RPA) reaction for the detection and identification of the high-risk HPV-16 genotype. The electrochemical biosensor exhibited a high degree of specificity and sensitivity, as evidenced by its limit of detection (LOD) of 0.23 copies/μL of HPV-16 DNA. The validity of this electrochemical platform was confirmed through the analysis of 40 cervical tissues samples, and the findings were consistent with those obtained through polymerase chain reaction (PCR) testing. Our straightforward electrochemical detection technology and quick turnaround time at 75 min make the assay suitable for point-of-care testing in low-resource settings.
Serum levels of dengue virus (DENV) non-structural 1 (NS1) antigen can serve as a valuable prognostic indicator of severe dengue infections. A quartz crystal microbalance (QCM)-based biosensor with a biomimetic recognition element was designed to quantitatively detect DENV NS1 as an early disease biomarker. To mitigate the reliance on costly viral antigens during the molecular imprinting process, a synthetic peptide mimicking a DENV NS1 epitope was used as a surrogate template for the synthesis of an epitope-imprinted polydopamine (EMIPDA) sensing film on the biosensor surface. The maximal frequency shift for DENV NS1 was obtained with an EMIPDA film synthesised using 5 mg mL-1 of dopamine monomer and 0.5 mg mL-1 of peptide template. The EMIPDA-QCM biosensor achieved low detection and quantitation limits of 0.091 μg mL-1 and 0.436 μg mL-1, respectively, allowing acute-phase detection of dengue and prognosis of the disease progression. The EMIPDA-QCM biosensor exhibited remarkable selectivity with up to 68-fold larger frequency responses towards DENV NS1 compared to a major serum protein. The site-specific imprinting approach not only enhanced the biosensing performance but also enabled a 26-fold cost reduction for biosensor functionalisation, providing a cost-effective strategy for label-free biosensing of the dengue biomarker via the biopolymer film.
In this work, we proposed a biosensor for trypsin proteolytic activity assay using immobilization of model peptides on screen-printed electrodes (SPE) modified with gold nanoparticles (AuNPs) prepared by electrosynthetic method. Sensing of proteolytic activity was based on electrochemical oxidation of tyrosine residues of peptides. We designed peptides containing N-terminal cysteine residue for immobilization on an SPE, modified with gold nanoparticles, trypsin-specific cleavage site and tyrosine residue as a redox label. The peptides were immobilized on SPE by formation of chemical bonds between mercapto groups of the N-terminal cysteine residues and AuNPs. After the incubation with trypsin, time-dependent cleavage of the immobilized peptides was observed by decline in tyrosine electrochemical oxidation signal. The kinetic parameters of trypsin, such as the catalytic constant (kcat), the Michaelis constant (KM) and the catalytic efficiency (kcat/KM), toward the CGGGRYR peptide were determined as 0.33 ± 0.01 min-1, 198 ± 24 nM and 0.0016 min-1 nM-1, respectively. Using the developed biosensor, we demonstrated the possibility of analysis of trypsin specificity toward the peptides with amino acid residues disrupting proteolysis. Further, we designed the peptides with proline or glutamic acid residues after the cleavage site (CGGRPYR and CGGREYR), and trypsin had reduced activity toward both of them according to the existing knowledge of the enzyme specificity. The developed biosensor system allows one to perform a comparative analysis of the protease steady-state kinetic parameters and specificity toward model peptides with different amino acid sequences.
A paper-based polymeric ionic liquid (p-Poly-(MMA-IL)) was successfully developed by grafting the polymeric ionic liquid on the surface of commercial filter paper (FP) by using the dipping method, presenting a new cost-effective film. The newly developed p-Poly-(MMA-IL) FP was then applied as a paper-based thin-film microextraction (p-TFME) analytical device to extract 14 compounds as representative of five groups of antibiotic drugs, which were sulfonamides, tetracyclines, fluoroquinolones, penicillin and macrolides in environmental water samples. Besides, p-Poly-(MMA-IL) FP, p-Poly-(MMA) FP, and unmodified filter paper were successfully characterised by FTIR, NMR, FESEM, TGA, and XRD techniques. They underwent significant parameters optimisation, which affected the extraction efficiency. Under optimal conditions, the proposed (p-Poly-(MMA-IL) FP-TFME) device method was evaluated and applied to analyse multi-class antibiotic drugs in environmental water samples by using a liquid chromatography-mass spectrometry (LC-MS). The validation method showed that a good linearity (0.1 μg L-1 - 500 μg L-1) was noted (R2 > 0.993, n = 3). Detection and quantification limits were within 0.05 μg L-1 - 4.52 μg L-1 and 0.15 μg L-1 - 13.6 μg L-1, respectively. The relative standard deviation (RSD) values ranged at 1.4%-12.2% (intra-day, n = 15) and 4.4%-11.0% (inter-day, n = 10). The extraction recoveries of environmental water samples ranged from 79.1% to 126.8%, with an RSD of less than 15.4% (n = 3). The newly developed paper-based polymeric ionic liquid (p-Poly-(MMA-IL) FP) for analysis of multi-class antibiotic drugs under the p-TFME analytical device procedure was successfully achieved with limited sample volume and organic solvent, fast extraction, and feasible in daily analysis. The detection concentration and relative RSD of multi-class antibiotics determined in various environmental water samples by the proposed method (n = 5) were within 0.44 μg L-1 - 54.41 μg L-1 and 0.69%-15.56%, respectively. These results signified the potential of the p-Poly-(MMA-IL) FP-TFME device as an efficient, sensitive and environmentally friendly approach for analysing antibiotics.
Oil and grease content in wastewater is used as an environmental monitoring parameter in the oil and gas industry to prevent serious pollution. Conventional oil and grease laboratory testing is time-consuming and necessitates the use of a hazardous chemical solvent, resulting in non-real-time test data and unnecessary chemical waste. On-site or real-time analysis can enable monitoring of oil and grease in wastewater before discharge to the environment from an operating plant, allowing immediate action to be taken to mitigate environmental impact before contamination spirals out of control. Bioluminescent whole-cell biosensors have been reported to have high sensitivity and selectivity in environmental samples, but only for a few traces of organic compounds such as polycyclic aromatics and naphthalene, allowing for faster analysis times. However, no evaluation of biosensor application for oil and grease (a mixture of hydrocarbons) detection in wastewater, which is critical in the oil and gas industry, has been published to date. Herein, the advantages, disadvantages, challenges, and limitations of using a whole-cell bioluminescent biosensor technology to measure oil and grease content in wastewater are carefully reviewed. This review attempts to bridge the knowledge gap between conventional laboratory methods and biosensor technology in terms of analytical challenges, identifying areas for improvement as well as real-world applications for oil and grease content detection in wastewater.
With each passing year, the agriculture and wood processing industries generate increasingly high tonnages of biomass waste, which instead of being burned or left to accumulate should be utilized more sustainably. In parallel, advances in green technology have encouraged large companies and nations to begin using eco-friendly materials, including eco-friendly emulsifiers, which are used in various industries and in bio-based materials. The emulsion-conducive properties of lignocellulosic materials such as cellulose, hemicellulose, and lignin, the building blocks of plant and wood structures, have demonstrated a particular ability to alter the landscape of emulsion technology. Beyond that, the further modification of their structure may improve emulsion stability, which often determines the performance of emulsions. Considering those trends, this review examines the performance of lignocellulosic materials after modification according to their stability, droplet size, and distribution by size, all of which suggest their outstanding potential as materials for emulsifying agents.
A dynamic pH junction was used in capillary electrophoresis (CE-DAD) to on-line preconcentrate, separate, and determine trace amounts of sulfonamide antibiotics (SAs) in milk and yoghurt samples in this study. A sample matrix with 0.15% acetic acid and 10% methanol (MeOH) at a pH of 4.0, and a background electrolyte (BGE) that contained 35 mM sodium citrate with 10% MeOH at a pH of 8.5, and an acidic barrage of 0.4% acetic acid with 10% MeOH at a pH of 2.5 were utilised to achieve a stacking effect for SAs through a dynamic pH junction. Under optimised conditions, the proposed preconcentration method showed good linearity (30-500 ng/mL, r2 ≥ 0.9940), low limits of detection (LODs) of 4.1-6.3 ng/mL, and acceptable analytes recovery (81.2-106.9%) with relative standard deviations (RSDs) within 5.3-13.7 (n = 9). The limits of quantification (LOQs) were below the maximum residue limit approved by the European Union (EU) in this type of matrices. Sensitivity enhancement factors of up to 129 were reached with the optimised dynamic pH junction using CE with a diode array detector (DAD). The method was used to determine SAs in fresh milk, low-fat milk, full-cream milk, and yoghurt samples.
Paper-based analytical devices (PADs) with four new designs could be fabricated using commercially available home-based scan-and-cut printer. They serve for miniaturised platforms for chemical analysis. Replication analysis of a sample together with the calibration (using the analyte standards at different concentrations) can be completed in a single run, by utilising smartphone as the detector. Some new approaches for choosing detection zones were suggested. The four proposed PAD designs here were used as models in microliter scale operation to demonstrate the well-known chemistries of colorimetric determinations of iron, phosphate, and hardness using 1,10-phenanthroline and simple aqueous guava leaf extract; molybdate, and EBT-EDTA complexometric titration, respectively, through calibrations: where Blue (B) value = 88.2log [Fe3+] - 80.8, R2 = 0.989; B value = 1.75 [Fe3+] + 0.198, R2 = 0.999; Grey scale (I) value = 1.77 [Fe3+] - 1.22, R2 = 0.997; Red (R) value = 16.1log [PO43-] + 8.95, R2 = 0.999; Hue (H) value = 43.3log [Ca2+] + 233, R2 = 0.994, respectively. For the hardness, using one of the PAD designs, true titration was also possible. Applications of the proposed devices and procedures were demonstrated for real world samples with validation. Additionally, kinetic study of the molybdenum blue for phosphate was demonstrated using one of the PADs.
Dot blot assays have always been associated with antibodies as the main molecular recognition element, which are widely employed in a myriad of diagnostic applications. With the rising of aptamers as the equivalent molecular recognition elements of antibodies, dot blot assays are also one of the diagnostic avenues that should be scrutinized for their amenability with aptamers as the potential surrogates of antibodies. In this review, the stepwise procedures of an aptamer-based dot blot assays are underscored before reviewing the existing aptamer-based dot blot assays developed so far. Most of the applications center on monitoring the progress of SELEX and as the validatory assays to assess the potency of aptamer candidates. For the purpose of diagnostics, the current effort is still languid and as such possible suggestions to galvanize the move to spur the aptamer-based dot blot assays to a point-of-care arena are discussed.
Fast and efficient separation remains a big challenge in high performance liquid chromatography (HPLC). The need for higher efficiency and resolution in separation is constantly in demand. To achieve that, columns developed are rapidly moving towards having smaller particle sizes and internal diameters (i.d.). However, these parameters will lead to high back-pressure in the system and will burden the pumps of the HPLC instrument. To address this limitation, monolithic columns, especially silica-based monolithic columns have been introduced. These columns are being widely investigated for fast and efficient separation of a wide range of molecules. The present article describes the current methods developed to enhance the column efficiency of particle packed columns and how silica monolithic columns can act as an alternative in overcoming the low permeability of particle packed columns. The fundamental processes behind the fabrication of the monolith including the starting materials and the silica sol-gel process will be discussed. Different monolith derivatization and end-capping processes will be further elaborated and followed by highlights of the performance such monolithic columns in key applications in different fields with various types of matrices.
COVID-19 pandemic is a serious global health issue today due to the rapid human to human transmission of SARS-CoV-2, a new type of coronavirus that causes fatal pneumonia. SARS -CoV-2 has a faster rate of transmission than other coronaviruses such as SARS and MERS and until now there are no approved specific drugs or vaccines for treatment. Thus, early diagnosis is crucial to prevent the extensive spread of the disease. The reverse transcription-polymerase chain reaction (RT-PCR) is the most routinely used method until now to detect SARS-CoV-2 infections. However, several other faster and accurate assays are being developed for the diagnosis of COVID-19 aiming to control the spread of infection through the identification of patients and immediate isolation. In this review, we will discuss the various detection methods of the SARS-CoV-2 virus including the recent developments in immunological assays, amplification techniques as well as biosensors.
Food contamination is a serious concern because of a high level of chemicals in food causes severe health issues. Safeguarding the public from the risk of adulterated foods has become a challenging mission. Chloropropanols are of importance to food safety and food security because they are common chemical food contaminants and believed to be carcinogenic to humans. In chemical sensing, chloropropanols are challenging analytes owing to the lacking diversity of functional groups and difficulty in targeting the hydroxyl group in aqueous environments. Moreover, because of their small molecular size, the compositions of chloropropanols remain challenging for achieving chromatographic determination. Herein, to simulate human smell and taste sensations, serum albumins, which are protein-based receptors, were introduced as low-selective receptors for differential sensing. Utilizing serum albumins, a fluorophore (PRODAN), and an additive (ascorbic acid), a differential-based optical biosensor array was developed to detect and differentiate chloropropanols. By integrating the sensor array with linear discriminant analysis (LDA), four chloropropanols were effectively differentiated based on their isomerism properties and the number of the hydroxyl groups, even at ultra-low concentration (5 nM). This concentration is far below the maximum tolerable level of 0.18 μM for chloropropanols. The sensing array was then employed for chloropropanols differentiation and quantification in the complex mixtures (e.g., synthetic soy and dark soy sauces). Leave-one-out cross-validation (LOOCV) analysis demonstrated 100% accurate classification for all tests. These results signify our differential sensing array as a practical and powerful tool to speedily identify, differentiate, and even quantify chloropropanols in food matrices.
The organophosphorous nerve agent VX is classified by the Chemical Warfare Convention (CWC) as a Schedule 1 chemical; namely a substance that is highly toxic with no use that is of benefit to society. Even with this classification, the nefarious use of the Schedule 1 chemical VX has been observed, as demonstrated in 2017 in Malaysia. Therefore, undertaking chemical analysis on samples of VX to identify chemical attribution signatures (CAS) for chemical forensics is required. To further understand the chemical profile of VX, and to aid in the identification of potential CAS, three in house synthesised stocks of VX were investigated. The three VX stocks analysed were synthesised in 2014, 2017 and 2018 using the same method, allowing for a comparison of data between each of the stocks at different stages of storage. As opposed to a majority of literature reports, these agent stocks were not stabilised, nor were they subjected to forced degradation. Using NMR, high resolution (HR) LC-HRMS, GC-(EI)MS and GC-(CI)MS to gain a full insight into the CAS profile, a total of 44 compounds were identified. Of these compounds, 30 were readily identified through accurate mass measurement and NIST library matches. A further seven were identified through extensive LC-HRMS/MS studies, with seven remaining unresolved. Several compounds, identified in minor amounts, were able to be traced back to impurities in the precursor compounds used in the synthesis of VX, and hence may be useful as CAS for source attribution.
The developments in mass spectrometry (MS) in the past few decades reveal the power and versatility of this technology. MS methods are utilized in routine analyses as well as research activities involving a broad range of analytes (elements and molecules) and countless matrices. However, manual MS analysis is gradually becoming a thing of the past. In this article, the available MS automation strategies are critically evaluated. Automation of analytical workflows culminating with MS detection encompasses involvement of automated operations in any of the steps related to sample handling/treatment before MS detection, sample introduction, MS data acquisition, and MS data processing. Automated MS workflows help to overcome the intrinsic limitations of MS methodology regarding reproducibility, throughput, and the expertise required to operate MS instruments. Such workflows often comprise automated off-line and on-line steps such as sampling, extraction, derivatization, and separation. The most common instrumental tools include autosamplers, multi-axis robots, flow injection systems, and lab-on-a-chip. Prototyping customized automated MS systems is a way to introduce non-standard automated features to MS workflows. The review highlights the enabling role of automated MS procedures in various sectors of academic research and industry. Examples include applications of automated MS workflows in bioscience, environmental studies, and exploration of the outer space.
An optical aptasensor-based sensing platform for rapid insulin detection was fabricated. Aminated porous silica microparticles (PSiMPs) were synthesized via a facile mini-emulsion method to provide large surface area for covalent immobilization of insulin-binding DNA aptamer (IGA3) by glutaraldehyde cross-linking protocol. A Nickel-salphen type complex with piperidine side chain [Ni(II)-SP] was synthesized with a simple one-pot reaction, and functionalized as an optical label due to strong π-π interaction between aromatic carbons of G-quadruplex DNA aptamer and planar aromatic groups of Ni(II)-SP to form the immobilized IGA3-Ni(II)-SP complex, i.e. the dye-labeled aptamer, thereby bringing yellow colouration to the immobilized G-quartet plane. Optical characterization of aptasensor towards insulin binding was carried out with a fiber optic reflectance spectrophotometer. The maximum reflectance intensity of the immobilized IGA3-Ni(II)-SP complex at 656 nm decreased upon binding with insulin as aptasensor changed to brownish orange colouration in the background. This allows optical detection of insulin as the colour change of aptasensor is dependent on the insulin concentration. The linear detection range of the aptasensor is obtained from 10 to 50 μIU mL-1 (R2 = 0.9757), which conformed to the normal fasting insulin levels in human with a limit of detection (LOD) at 3.71 μIU mL-1. The aptasensor showed fast response time of 40 min and long shelf life stability of >3 weeks. Insulin detection using healthy human serums with informed consent provided by participants suggests the DNA aptamer biosensor was in good agreement with ELISA standard method using BIOMATIK Human INS (Insulin) ELISA Kit.
As a widely consumed beverage, coffee tends to be a target for intentional adulteration. This study describes the application of modified quick, easy, cheap, effective, rugged, and safe (QuEChERS) coupled to liquid chromatography-high-resolution mass spectrometry (LC-HRMS) for simultaneous screening, identification, and quantification of undeclared phosphodiesterase 5 (PDE5) inhibitors in instant coffee premixes (ICPs). The mass spectrometer was operated in auto MS/MS acquisition for simultaneous MS and MS/MS experiments. Qualitative establishments from the suspected-target screening and targeted identification processes led to an unambiguous analyte assignment from the protonated molecule ([M+H]+) precursor ion which is subsequently used for quantification of 23 targeted PDE5 inhibitors. The analytical method validation covered specificity, linearity, range, accuracy, limit of detection (LOD), limit of quantification (LOQ), precisions, matrix effect (ME), and extraction recovery (RE). The specificity was established using the optimised chromatographic separation as well as the distinguishable [M+H]+ precursor ion. The linearity of each target analyte was demonstrated with a coefficient of determination (r2) of >0.9960 over the expected range of sample concentrations. The accuracy ranged from 88.1%-119.3% with LOD and LOQ of <70 ng/mL and 80 ng/mL, respectively. Excellent precisions were established within 0.4%-9.1% of the relative standard deviation. An insignificant ME within -5.2% to +8.7% was achieved using three different strategies of chromatography, sample extraction, and sample dilution. The RE was good for all target analytes within 84.7%-123.5% except for N-desethylacetildenafil at low (53.8%) and medium (65.1%) quality control levels. The method was successfully applied to 25 samples of ICPs where 17 of them were found to be adulterated with PDE5 inhibitors and their analogues. Further quantification revealed the total amount of these adulterants ranged from 2.77 to 121.64 mg per sachet.
Aptamers are nucleic acid-based molecular recognition elements that are specific and have high binding affinity against their respective targets. On account of their target recognition capacity, aptamers are widely utilized in a number of applications including diagnostics. This review aims to highlight the recent developments of aptasensors expedient for point-of-care (POC) diagnostics. Significant focus is given on the primary assay formats of aptamers such as fluorescence, electrochemical, surface plasmon resonance (SPR) and colorimetric assays. A potpourri of platforms such as paper-based device, lateral flow assay, portable electrodes, portable SPR and smart phones expedient for point-of-care (POC) diagnostics are discussed. Emphasis is also given on the technicalities and assay configurations associated with the sensors.
Molecularly imprinted silica gel (MISG) was incorporated through dispersion in agarose polymer matrix to form a mixed matrix membrane (MMM) and was applied for the determination of three sulfonamide antibiotic compounds (i.e. sulfamethoxazole (SMX), sulfamonomethoxine (SMM), and sulfadiazine (SDZ)) from environmental water samples. Several important microextraction conditions, such as type of desorption solvent, extraction time, amount of sorbent, sample volume, pH, and effect of desorption time, were comprehensively optimized. A preconcentration factors of ≥ 20 was achieved by the extraction of 12.5 mL of water samples using the developed method. This microextraction-HPLC method demonstrated good linearity (1-500 μg L-1) with a coefficient of determination (R2) of 0.9959-0.9999, low limits of detection (0.06-0.17 μg L-1) and limits of quantification (0.20-0.56 μg L-1), good analyte recoveries (80-96%), and acceptable relative standard deviations (< 10%) under the optimized conditions. The method is systematically compared to those reported in the literature.