On-site Identification Research Articles

Miniature lab-made electrochemical biosensor: A promising sensing kit for rapid detection of E. coli in water, urine and milk

A novel, rapid production methodology for laboratory-made carbon electrodes (LCE) employing cost-effective and readily available materials has developed in the present work. The LCE presents superior electrochemical characteristics compared to commercially available screen-printed carbon electrodes (SPCE). Furthermore, this research has demonstrated the performance of readily accessible, highly sensitive, and portable biosensors for on-site detection of E. coli in aqueous samples. Silver nanoparticles (AgNPs) were successfully electrodeposited onto the LCE (Ag-LCE) using the electrochemical method at optimised parameters. The E. coli-specific aptamer was conjugated with AgNPs, and uncoated Ag-LCE surfaces were blocked with a BSA (BSA-Apt-Ag-LCE). The developed BSA-Apt-Ag-LCE biosensor was characterised and validated for the successful detection of E. coli in aqueous samples using cyclic voltammetry (CV). A linear correlation was obtained for sensor response in the 3.4 × 101 to 3.4 × 106 CFU/ml bacterial concentration as ΔIpa = 5.71 log C + 2.91 with R2 = 0.987. BSA-Apt-Ag-LCE biosensors have a limit of detection of 34 CFU/ml and a response time of 15 min, indicating their prompt and practical on-site identification capabilities. The proficient detection of E. coli in diverse aqueous samples, substantiated by its consistent reproducibility as indicated by the relative standard deviation (RSD) value of a maximum of 1.71 %, is a compelling validation of the biosensor's efficacy and reliability. The proposed biosensor exhibited selectivity towards E. coli and was found stable even after being stored at 4 °C for four weeks.

A LAMP-based colorimetric and fluorescence dual-channel assay for on-site identification of adulterated meat by a portable device

Rapid on-site identification of meat adulteration is essential for ensuring food safety. The Loop-mediated isothermal amplification (LAMP) technique is an ideal solution for on-site diagnosis due to its simplicity, speed, high specificity, and sensitivity. In this study, we designed pork-specific primers for LAMP and optimized the LAMP reaction conditions. By incorporating hydroxynaphthol blue and calcein indicators into the reaction system, the LAMP results can be visually observed without the need for specialized equipment. We also engineered a self-designed, portable device that integrates LAMP colorimetric and fluorescence detection, termed LAMP-PD, to achieve both amplification and dual-channel detection. This device is cost-effective, priced at only $54.65. The detection results from LAMP-PD were found to be comparable to those of commercial spectrophotometers, confirming its accuracy. The LAMP-PD is capable of detecting pork at concentrations as low as 0.1% in adulterated meat samples, highlighting the high sensitivity of the LAMP assay. The portability and affordability of the LAMP-PD underscore its significant potential for on-site identification of adulterated meat samples.

Electrochemical Impedance Spectroscopy-Based Microfluidic Biosensor Using Cell-Imprinted Polymers for Bacteria Detection.

The rapid and sensitive detection of bacterial contaminants using low-cost and portable point-of-need (PoN) biosensors has gained significant interest in water quality monitoring. Cell-imprinted polymers (CIPs) are emerging as effective and inexpensive materials for bacterial detection as they provide specific binding sites designed to capture whole bacterial cells, especially when integrated into PoN microfluidic devices. However, improving the sensitivity and detection limits of these sensors remains challenging. In this study, we integrated CIP-functionalized stainless steel microwires (CIP-MWs) into a microfluidic device for the impedimetric detection of E. coli bacteria. The sensor featured two parallel microchannels with three-electrode configurations that allowed simultaneous control and electrochemical impedance spectroscopy (EIS) measurements. A CIP-MW and a non-imprinted polymer (NIP)-MW suspended perpendicular to the microchannels served as the working electrodes in the test and control channels, respectively. Electrochemical spectra were fitted with equivalent electrical circuits, and the charge transfer resistances of both cells were measured before and after incubation with target bacteria. The charge transfer resistance of the CIP-MWs after 30 min of incubation with bacteria was increased. By normalizing the change in charge transfer resistance and analyzing the dose-response curve for bacterial concentrations ranging from 0 to 107 CFU/mL, we determined the limits of detection and quantification as 2 × 102 CFU/mL and 1.4 × 104 CFU/mL, respectively. The sensor demonstrated a dynamic range of 102 to 107 CFU/mL, where bacterial counts were statistically distinguishable. The proposed sensor offers a sensitive, cost-effective, durable, and rapid solution for on-site identification of waterborne pathogens.

The Time Is Ripe: Olive Drupe Maturation Can Be Simply Evidenced by a Miniaturized, Portable and Easy-to-Use MicroNIR Green Sensor

The analytical study described in this work, based on NIR spectroscopy with a handheld device, allowed the development of a chemometric prediction model that has been validated for the objective evaluation of the ripening of olive drupes. The miniaturized, portable NIR spectrometer is proposed here as an easy-to-use sensor able to estimate the best harvesting time for ripening of olive drupes. The MicroNIR/chemometrics approach was developed for on-site identification of olive drupe ripening directly on plants, avoiding collection and successive laboratory analysis steps. A supporting parallel characterization by chromatographic techniques validated the spectroscopic prediction. The novelty of this approach consists in the possibility of investigating the olive drupe maturation point by collecting spectra in the near-infrared region and processing them using a chemometric model. The fast and accurate device allows one to easily follow the spectrum profile changes of olive drupes during ripening, thus preserving the fruits from being harvested too early or too late. The results of this study demonstrate the possibility of using the MicroNIR/chemometrics approach to determine the optimal ripening time of olives regardless of the plant variety, age and cultivation location. The results consequently demonstrated that the MicroNIR/chemometrics approach can be proposed as a new method to perform on-site evaluation of ripening by a single-click device. It can be conveniently used by any operator, who does not necessarily have to be expert but must simply be trained to use spectroscopy and a prediction model.

Development and evaluation of rapid and simple detection of Klebsiella pneumoniae using closed dumbbell-mediated isothermal amplification diagnostic assay.

Klebsiella pneumoniae (K. pneumoniae) is the most common pathogen causing hospital respiratory tract infection and epidemic. Gold standard procedures of microscopic examination and biochemical identification are widely used in clinical diagnosis with disadvantages of low sensitivity, time-consuming and sophisticated equipment requiring. An efficient, nucleic acid amplification-based sensitive and specific on-site identification of K. pneumoniae in clinical is necessary to facilitate clinical medication and disease control. We developed a closed dumbbell mediated isothermal amplification (CDA) assay for the rapid and sensitive detection of conserved rcsA gene in K. pneumoniae by real-time fluorescence monitoring and end-point colorimetric judgement. We designed and selected a pair of inner primers of CDA to detect K. pneumoniae. Then outer and loop primers were designed and verified to accelerate CDA reaction to achieve more efficient detection of K. pneumoniae. The results showed the detection limit of CDA assay was 1.2 × 10-5 ng/μL (approximately 1 copy of the target gene) within 60 min, which was 100-fold more sensitive than real-time quantitative PCR (qPCR). Several pathogen genomic DNAs (Staphylococcus aureus, Shigella sonnei, Vibrio parahaemolyticus, Escherichia coli, Candida glabrata, Candida tropicalis, Candida parapsilosis, Candida albicans, Streptococcus agalactiae, Rickettsia, Listeria monocytogenes, Pseudomonas aeruginosa, Klebsiella oxytoca, and Klebsiella aerogenes) were used to evaluate the sensitivity and specificity of the established K. pneumoniae CDA assay. Total 224 batches of samples from other strains tested were negative and 296 batches of extracted K. pneumoniae DNA samples were positive by the developed CDA amplification approach, revealing high specificity and specificity of the diagnostic assay. In addition, the results of real-time fluorescence amplification of the K. pneumoniae CDA were in consistent with those of end-point colorimetric results. The established real-time fluorescence and visual CDA assays of K. pneumoniae with merits of rapid, sensitive and specificity could be helpful for on-site diagnosis and clinical screening in rural areas.

A ratiometric, turn-on fluorosensor for specific detection of Zn2+ ions

A potentially active fluorescent chemosensor (E)-2-(((2-(1H-benzo[d]imidazol-2-yl)phenyl)imino)methyl)-5-(diethylamino)phenol (BMDP) is developed by the condensation of 2-(2-aminophenyl)-1H-benzimidazole and 4-(diethylamino)salicylaldehyde. It shows an exceptional ratiometric fluorescence behavior when exposed to Zn2+ ions and the photoluminescence color is changed from blue to cyan under the exposure of a 365 nm UV light which is also strongly evident from the color chromaticity diagram. The exceptional ratiometric fluorescence behavior in the presence of Zn2+ ions is found to be due to the chelation-enhanced fluorescence (CHEF). The 1:1 stoichiometry of BMDP and Zn2+ ions in Zn2+ ions chelated BMDP complex is confirmed through Job’s plot analysis. Remarkably, Cd2+ does not interfere with the selective ratiometric fluorescence behavior induced by Zn2+ ions. The lowest detection limit and quantification limit are estimated to be 28 nM and 92 nM respectively. The cyan fluorescence of the Zn2+ ions chelated BMDP complex is interestingly reversed to the original BMDP in the presence of ethylenediamine tetraacetic acid (EDTA). At the molecular level, this can be thought of as a complimentary “INHIBIT” logic gate. Furthermore, it is possible to utilize the probe for on-site identification in test strips with improved Zn2+ ions selectivity. We have also used a smartphone-based method to demonstrate the usefulness of BMDP for the immediate quantification and detection of Zn2+ ions. In light of this, probe BMDP is a trustworthy fluorescence probe for on-site monitoring that can accurately identify Zn2+ ions even when some other metals and anions compete.

Duplex recombinase polymerase amplification combined with CRISPR/Cas12a-Cas12a assay for on-site identification of yak meat adulteration

Yak meat is known for its unique flavor and high nutritional value. Its low output results in high prices, and as a result, food fraud is often committed in the marketplace by using low-value cattle meat with similar morphological characteristics to pass off as high-value yak meat. However, the current detection methods for yak meat authenticity are mostly confined to laboratories and suffer from expensive equipment, high professional requirements, and insufficient specificity, making it difficult to realize rapid on-site assay of yak meat authenticity. In this study, a duplex recombinase polymerase amplification combined with CRISPR/Cas12a-Cas12a assay strategy for rapid visual on-site identification of cattle- and yak-derived components was established. With only a portable box containing some cheap and small equipment, the method can accurately identify the authenticity of yak meat products within 1 h. It has been verified that to this method’s detection limit can reach to 1 % (w/w) at least for the cattle meat, regardless of raw or cooked products. After testing the blind samples, 52.8 % of the yak meat products were adulterated, which was consistent with the results of the real-time PCR method, proving that the method has good accuracy and practicability. It can be used to achieve accuracy, rapid and on-site assay of the authenticity of various yak meat products.

Solid-state brightness and Al3+ ions-triggered flower-shaped nano-luminogen for cascade detection of Al3+ and PO43− ions

Solid-state photoluminescence materials have come out as an emergent material for their versatile optoelectronic applications. In the present contribution, a novel fluorescent probe, HMNS has been introduced which exhibits bright orange color photoluminescence when exposed to a 365 nm UV lamp and with a few drops of dimethyl sulfoxide (DMSO), it can be used as photoluminescence security ink. Further, HMNS is utilized for quickly and accurately detecting Al3+ ions in a water-DMSO (1:1, v/v) media displaying Al3+ ions-triggered aggregation-induced emission (AIE). Scanning electron microscopy and dynamics light scattering investigations revealed that HMNS from Al3+ ions triggered flower-shaped nano-luminogen. Probe HMNS can detect Al3+ ions within μM range and also the probe binds with Al3+ ions under physiological pH. The reaction time to Al3+ ions is only a few minutes (around 6–7 min). The development of J-aggregates and chelation-enhanced fluorescence (CHEF) in the water-DMSO (1:1, v/v) media may be responsible for the AIE behavior. Additionally, using the Al3+ ions triggered HMNS nano-luminogen could be used as a "marker-ink" which could be used as printing material to decipher secret data under UV light. Also, to determine its reversibility and reusability, the tested paper was washed with a PO43− ions solution. A notable decrease in the code's intensity indicates that this material can be used repeatedly. Moreover, the probe is suitable for on-site identification when employed in test strips with better Al3+ ions selectivity. We have also used a smartphone-based method to demonstrate the usefulness of HMNS for the immediate quantification and detection of Al3+ ions. In light of this, probe HMNS is a trustworthy on-site monitoring fluorescence probe that can accurately identify Al3+ and PO43− ions successively even when some other metals and anions compete.

Advanced nutrient monitoring: Cost-effective cerium ion detection with smart device integration

This research is critically important for large-scale poultry farming, where vigilant monitoring is necessary to assess nutrient levels in poultry feed. Detecting cerium in poultry feed through colorimetric techniques is crucial for evaluating nutrient intake, feed quality, animal health, and broader environmental impacts. This study significantly contributes to ongoing research and monitoring efforts, aiding in maintaining an optimal mineral balance and enhancing chicken quality and overall poultry performance. The study utilized Alizarin Red dye (AR), both individually and in combination with Eriochrome Black T (EBT), to identify the presence of cerium ions. Various experimental conditions such as pH, concentration, temperature, and specificity were thoroughly examined. The UV-Vis absorption spectra indicated that the average minimum detectable limit for cerium ions is approximately 5 ppm, with a detection range of 0.2–3 mM. Additionally, a cost-effective paper-based sensor and a portable colorimetric method for detecting cerium ions were innovatively designed. This paper-based sensor ensures precise detection at room temperature, demonstrating high sensitivity and selectivity. The detection system was integrated with smart devices, enabling the swift capture of Red, Green, and Blue (RGB) values for practical real-time applications. This integration allows for rapid on-site identification of cerium ions. The introduction of this affordable, accurate, and portable colorimetric method for monitoring cerium ions represents a promising advancement in developing accessible tools within this field.

Assessment of a Single Quadrupole Mass Spectrometer Combined with an Atmospheric Solids Analysis Probe for the On-Site Identification of Amnesty Bin Drugs.

The surging number of people who abuse drugs has a great impact on healthcare and law enforcement systems. Amnesty bin drug analysis helps monitor the "street drug market" and tailor the harm reduction advice. Therefore, rapid and accurate drug analysis methods are crucial for on-site work. An analytical method for the rapid identification of five commonly detected drugs ((3,4-methylenedioxymethamphetamine (MDMA), cocaine, ketamine, 4-bromo-2,5-dimethoxyphenethylamine, and chloromethcathinone)) at various summer festivals in the U.K. was developed and validated employing a single quadrupole mass spectrometer combined with an atmospheric pressure solids analysis probe (ASAP-MS). The results were confirmed on a benchtop gas chromatography-mass spectrometry instrument and included all samples that challenged the conventional spectroscopic techniques routinely employed on-site. Although the selectivity/specificity step of the validation assessment of the MS system proved a challenge, it still produced 93% (N = 279) and 92.5% (N = 87) correct results when tested on- and off-site, respectively. A few "partly correct" results showed some discrepancies between the results, with the MS-only unit missing some low intensity active ingredients (N-ethylpentylone, MDMA) and cutting agents (caffeine, paracetamol, and benzocaine) or detecting some when not present. The incorrect results were mainly based on library coverage. The study proved that the ASAP-MS instrument can successfully complement the spectroscopic techniques used for qualitative drug analysis on- and off-site. Although the validation testing highlighted some areas for improvement concerning selectivity/specificity for structurally similar compounds, this method has the potential to be used in trend monitoring and harm reduction.

Universal CRISPR-Cas12a and Toehold RNA Cascade Reaction on Paper Substrate for Visual Salmonella Genome Detection.

Salmonella, the most prevalent food-borne pathogen, poses significant medical and economic threats. Swift and accurate on-site identification and serotyping of Salmonella is crucial to curb its spread and contamination. Here, a synthetic biology cascade reaction is presented on a paper substrate using CRISPR-Cas12a and recombinase polymerase amplification (RPA), enabling the programming of a standard toehold RNA switch for a genome of choice. This approach employs just one toehold RNA switch design to differentiate between two different Salmonella serotypes, i.e., S. Typhimurium and S. Enteritidis, without the need for reengineering the toehold RNA switch. The sensor exhibits high sensitivity, capable of visually detecting as few as 100 copies of the whole genome from a model Salmonella pathogen on a paper substrate. Furthermore, this robust assay is successfully applied to detect whole genomes in contaminated milk and lettuce samples, demonstrating its potential in real sample analysis. Due to its versatility and practical features, genomes from different organisms can be detected by merely changing a single RNA element in this universal cell-free cascade reaction.

Detection and Identification of Pesticides in Fruits Coupling to an Au-Au Nanorod Array SERS Substrate and RF-1D-CNN Model Analysis.

In this research, a method was developed for fabricating Au-Au nanorod array substrates through the deposition of large-area Au nanostructures on an Au nanorod array using a galvanic cell reaction. The incorporation of a granular structure enhanced both the number and intensity of surface-enhanced Raman scattering (SERS) hot spots on the substrate, thereby elevating the SERS performance beyond that of substrates composed solely of an Au nanorod. Calculations using the finite difference time domain method confirmed the generation of a strong electromagnetic field around the nanoparticles. Motivated by the electromotive force, Au ions in the chloroauric acid solution were reduced to form nanostructures on the nanorod array. The size and distribution density of these granular nanostructures could be modulated by varying the reaction time and the concentration of chloroauric acid. The resulting Au-Au nanorod array substrate exhibited an active, uniform, and reproducible SERS effect. With 1,2-bis(4-pyridyl)ethylene as the probe molecule, the detection sensitivity of the Au-Au nanorod array substrate was enhanced to 10-11 M, improving by five orders of magnitude over the substrate consisting only of an Au nanorod array. For a practical application, this substrate was utilized for the detection of pesticides, including thiram, thiabendazole, carbendazim, and phosmet, within the concentration range of 10-4 to 5 × 10-7 M. An analytical model combining a random forest and a one-dimensional convolutional neural network, referring to the important variable-one-dimensional convolutional neural network model, was developed for the precise identification of thiram. This approach demonstrated significant potential for biochemical sensing and rapid on-site identification.

Intelligent detection of tetracycline by a rare earth multicolor fluorescent probe based on guanosine-5′-monophosphate

Long-term discharge of tetracycline (TC) in the environment exerts selective pressure on the bacteria, increasing resistance in sensitive bacteria and posing a serious threat to public health. Therefore, it is crucial to develop real-time detection of tetracycline. In this study, we reported a novel rare-earth fluorescent nanosensor FITC-GMP-Eu, based on the formation of guanosine-5′-monophosphate disodium salt (GMP) and fluorescein isothiocyanate (FITC), which enabled the real-time identification of tetracycline in the environment. When there was no TC in the environmental sample, the FITC-GMP-Eu nanosensor exhibited the green fluorescence of FITC due to the fluorescence quenching effect of water molecules on Eu3+. When the environmental samples contained TC, the red fluorescence of Eu3+ could be sensitized due to the diketone structure in the tetracycline molecule, which enabled the naked-eye visualized identification of TC. The nanosensor had high sensitivity (detection limit of 19 nM) and a wide detection range (0–30 μM) for TC. At the same time, the sensor was characterized by high visualization and anti-interference ability due to the strong fluorescence of both FITC and Eu complexes. In order to improve the practicality of the detection, this study also prepared a paper-based and sodium alginate gelated fluorescence detection platform with low background fluorescence. Combined with the common color analysis and RGB extraction procedures in mobile phones, real-time and on-site identification of TC on the surface of various fruit samples can be achieved.

Target-Responsive DNA Nanoclaw for the On-Site Identification of Chinese Medicines with Naked Eye.

The identification of Chinese medicinal herbs occupies a crucial part in the development of the food and drug market. Although molecular identification based on real-time PCR offers good versatility and uniform digital standards compared with traditional methods, such as morphology, the dependence on large-scale equipment hinders spot detection and marketable applications. In this study, we developed a DNA nanoclaw for colorimetric detection and visible on-site identification of Chinese medicines. When specific miRNA is present, the DNAzyme is activated and cleaves the substrate strand, triggering the catalytic hairpin assembly (CHA) reaction and forming branched DNA junctions on AuNP-I. This can then capture AuNP-II through hybridization and facilitate their aggregation, resulting in a noticeable color change that is observable to the naked eye. By harnessing the dual amplification of DNAzyme and CHA, this highly sensitive nanoprobe successfully achieved specific identification of Chinese medicines. This offers a new perspective for on-site testing in the herbal market.

Analysis of Whole-Genome facilitates rapid and precise identification of fungal species.

Fungal identification is a cornerstone of fungal research, yet traditional molecular methods struggle with rapid and accurate onsite identification, especially for closely related species. To tackle this challenge, we introduce a universal identification method called Analysis of whole GEnome (AGE). AGE includes two key steps: bioinformatics analysis and experimental practice. Bioinformatics analysis screens candidate target sequences named Targets within the genome of the fungal species and determines specific Targets by comparing them with the genomes of other species. Then, experimental practice using sequencing or non-sequencing technologies would confirm the results of bioinformatics analysis. Accordingly, AGE obtained more than 1,000,000 qualified Targets for each of the 13 fungal species within the phyla Ascomycota and Basidiomycota. Next, the sequencing and genome editing system validated the ultra-specific performance of the specific Targets; especially noteworthy is the first-time demonstration of the identification potential of sequences from unannotated genomic regions. Furthermore, by combining rapid isothermal amplification and phosphorothioate-modified primers with the option of an instrument-free visual fluorescence method, AGE can achieve qualitative species identification within 30 min using a single-tube test. More importantly, AGE holds significant potential for identifying closely related species and differentiating traditional Chinese medicines from their adulterants, especially in the precise detection of contaminants. In summary, AGE opens the door for the development of whole-genome-based fungal species identification while also providing guidance for its application in plant and animal kingdoms.

Nanoparticles Enhanced Laser-Induced Breakdown Spectroscopy for Characterisation and Discrimination of Gemstones

Laser-induced breakdown spectroscopy (LIBS) is an excellent technique for rapid on-site investigations that attracts interest from diverse research areas. Gemmology is no exception. The application of LIBS for gemstone characterisation is limited due to ineffective ablation and crack formation, even more so with cost-effective, non-gated LIBS systems. Nanoparticle-enhanced LIBS (NELIBS) has proven to improve the effectiveness of LIBS by minimizing sample damage and enhancing the spectral features. Therefore, this study is dedicated to exploring the advantages of NELIBS, for characterising Sapphire and Opal and discrimination based on spectral differences. Our objective is to explore enhancing spectral features and performing discriminant analysis using the PLS-DA algorithm. Nanoparticles (NPs) were deposited in two layers by sequentially drying two drops (2µL) of a colloidal solution of 20nm gold nanoparticles (AuNPs) on the sample surface. Targeted areas were shot with 3 pulses of Nd:YAG laser (~350mJ, 10ns, 1064nm, 1Hz) for collecting NELIBS spectra with the OceanOptics HR4000 spectrometer. The procedure was repeated without NPs for comparative analysis with conventional LIBS (CLIBS). Results have shown a significant enhancement in spectral features, i.e., the emergence of several new spectral lines of major gemstone elements in the UV-Vis regions of the NELIBS spectra, while the CLIBS spectra were devoid of any meaningful spectral information. The PLS-DA model was trained and validated using a 4fold cross-validation approach. The model discriminated gemstones with 99.48% accuracy at the 4th fold and exhibited a mean cross-validation accuracy of 98.97%. This preliminary investigation demonstrates the effectiveness of NELIBS for characterization and the potential for onsite identification of gemstones.

A point-of-care testing platform for on-site identification of genetically modified crops.

Genetically modified (GM) food is still highly controversial nowadays. Due to the disparate policies and attitudes worldwide, demands for a rapid, cost-effective and user-friendly GM crop identification method are increasingly significant for import administration, market supervision, etc. However, as the most-recognized methods, nucleic acid-based identification approaches require bulky instruments, long turn-around times and trained personnel, which are only suitable in laboratories. To fulfil the urgent needs of on-site testing, we develop a point-of-care testing platform that is able to identify 12 types of GM crops in less than 40 minutes without using laboratory settings. Our system integrates sample pre-treatment modules in a microfluidic chip, performs DNA amplification via a battery-powered portable kit, and presents results via eye-recognized colorimetric change. A paraffin-based reflow method and a slip plate-based fluid switch are developed to encapsulate and release amplification primers in individual microwells on demand, thus enabling identification of varied targets simultaneously. Our system offers an efficient, affordable and convenient tool for GM crop identification, thus it will not only benefit customs and market administration bureaus, but also satisfy demands of numerous consumers.

Rapid RNase H-dependent PCR lateral flow assay for the detection of red snapper

Red snapper (Lutjanus campechanus or Lutjanus purpureus) is one of the most commercially important and commonly mislabeled seafood species in the United States. Existing molecular methods for species identification are expensive and require being sent to an off-site laboratory, which takes multiple days to process. In this study, we have standardized a rapid rhPCR-coupled lateral flow assay for the identification of red snapper species. Our assay involved a simple heat-lysis DNA extraction procedure and a multiplex PCR reaction, consisting of a red snapper single nucleotide polymorphism (SNP)-specific rhPCR primer, thermotolerant RNase H2 enzyme, and an internal amplification control (IAC) for their onsite identification. Amplicons generated in the PCR reaction were detected using dual target lateral flow strips. The standardized assay was validated with 108 barcoded fish samples. Samples identified as Lutjanus campechanus or L. purpureus by DNA barcoding formed three distinct bands, while other fish species formed only two bands on the lateral flow strips. A minimum of 0.37 ng of crude DNA was detectable in the rhPCR reaction and generated a visible band on the lateral flow dipstick. The assay showed 100% specificity and took 90–120 min to complete. The assay generated results that can be used to authenticate red snapper at the wholesale and retail levels of seafood processing and commerce. Our results confirm the ability of rhPCR-lateral flow assays to be an alternative tool for onsite species authentication for the seafood industry.

Analysis of the fluorescent properties of vaginal fluid upon ageing

Detection and identification of body fluids are crucial aspects of forensic investigations, aiding in crime scene reconstructions and providing important leads. Although many methods have been developed for these purposes, no method is currently in use in the forensic field that allows rapid, non-contact detection and identification of vaginal fluids directly at the crime scene. The development of such technique is mainly challenged by the complex chemistry of the constituents, which can differ between donors and exhibits changes based on woman’s menstrual cycle. The use of fluorescence spectroscopy has shown promise in this area for other biological fluids. Therefore, the aim of this study was to identify specific fluorescent signatures of vaginal fluid with fluorescence spectroscopy to allow on-site identification. Additionally, the fluorescent properties were monitored over time to gain insight in the temporal changes of the fluorescent spectra of vaginal fluid. The samples were excited at wavelengths ranging from 200 to 600 nm and the induced fluorescence emission was measured from 220 to 700 nm. Excitation and emission maps (EEMs) were constructed for eight donors at seven time points after donation. Four distinctive fluorescence peaks could be identified in the EEMs, indicating the presence of proteins, fluorescent oxidation products (FOX), and an unidentified component as the dominant contributors to the fluorescence. To further asses the fluorescence characteristics of vaginal fluid, the fluorescent signatures of protein and FOX were used to monitor protein and lipid oxidation reactions over time. The results of this study provide insights into the intrinsic fluorescent properties of vaginal fluid over time which could be used for the development of a detection and identification method for vaginal fluids. Furthermore, the observed changes in fluorescence signatures over time could be utilized to establish an accurate ageing model.

Simultaneous Detection of Citric Acid and Oxalic Acid Based on Dual Spectrum and Biomimetic Peroxidase for Urolithiasis Screening with a Fully Automatic Urine Analyzer.

Urolithiasis stands as a prevalent ailment within the urinary system, with hyperoxaluria and hypocitraturia being the most frequent manifestations characterized by excessive oxalic acid (OA) and deficient citric acid (CA) levels in urine. Detecting these compounds in urine quantitatively holds paramount importance for early urolithiasis screening. Existing methodologies fall short in achieving simultaneous and on-site identification of OA and CA, posing challenges for accurate urolithiasis screening. Addressing this concern, the study successfully accomplishes the concurrent identification of OA and CA in urine through a combination of dual-spectral analysis and biomimetic peroxidase utilization. Bovine serum albumin and dithiothreitol-modified copper nanoclusters (BSA-DTT-CuNCs) are employed as biomimetic peroxidases, effectively mitigating interference and enabling the simultaneous determination of OA and CA. The quantification range spans from 0 to 12mm for OA and 0.5 to 2.5mm for CA, with detection limits of 0.18 and 0.11mm, respectively. To facilitate swift and on-location urine analysis, a fully automated urine analyzer (FAUA) is introduced that streamlines the process of biomarker pretreatment and identification within urine samples. Validation with real urine samples from urolithiasis patients demonstrates the method's diagnostic precision, highlighting the dual-spectral technique and analyzer's promising role in urolithiasis screening.