Femtogram Level Research Articles

Rapid Detection of a Downy Mildew Pathogen, Peronospora destructor, in Infected Onion Tissues and Soils by Loop-Mediated Isothermal Amplification.

Downy mildew of onion caused by a soil-inhabiting water mold, Peronospora destructor, is one of the most devastating diseases that can destroy entire onion fields in a matter of days. In this study, we developed a loop-mediated isothermal amplification (LAMP) assay that allows for rapid detection of P. destructor by visual inspection. The internal transcribed spacer 2 region of P. destructor was used to design primer sets for LAMP reactions. The optimal temperature and incubation time were determined for the most efficient primer set. In the optimized condition, the LAMP assay exhibited at least 100 times more sensitivity than conventional PCR, detecting femtogram levels of P. destructor genomic DNA (gDNA). Detection of the pathogen from a small number of spores without gDNA extraction further confirmed the high sensitivity of the assay. For specificity, the LAMP assay was negative for gDNA of other fungal pathogens that cause various diseases on onion and oomycetes, whereas the assay was positive for gDNA extracted from onion tissues showing the typical downy mildew symptoms. Finally, we examined the efficacy of the LAMP assay in detection of P. destructor in soils. Soils collected from onion fields that had been contaminated with P. destructor were solarized for 60 days. Whereas the LAMP assay was negative for the solarized soils, we were able to detect P. destructor that oversummers in fields. The LAMP assay developed in this study enables rapid detection and diagnosis of downy mildew of onion in infected tissues and in soil.

Highly sensitive in-situ growth gold dendrite structure electrochemical sensor for early Alzheimer's disease screening

Beta-amyloid 42 (Aβ-42) peptide is an essential biomarker for Alzheimer's disease (AD) diagnosis. This study presents high sensitivity and stability sensors for Aβ-42 peptide detection, along with an integrated handheld analysis and display device. The sensors, with a wide response range and high accuracy, capable of measuring Aβ-42 peptide concentration at the femtogram level, could have enabled early screening for AD. Employing an in-situ grown three-dimensional sensor, the specific surface area of the sensor was increased by 200 % which, thereby, provided more antibody binding sites to enhance detection accuracy. This electrochemical sensor demonstrated a broad linear response range from 4.5 × 10−8 pg/mL to 4.5 × 102 pg/mL and could accurately determine the concentration of spiked analytes (including artificial saliva, human plasma, and human blood). Furthermore, the sensor exhibited satisfactory stability, with only a 5 % sensitivity loss after 35 days of storage at 4 °C. Additionally, this handheld Aβ-42 peptide testing device incorporates a display for result delivery. Such innovative technology could fill the gap in the market for portable testing instruments and could provide an efficient and reliable solution for early AD screening.

Simultaneous Determination of Transuranium Radionuclides in Urine by Tandem Quadrupole ICP-MS/MS with Mass-Shift Mode Combined with Chemical Separation.

The urine bioassay method for transuranium nuclides (237Np, 239,240,241Pu, 241Am, and 244Cm) is needed to quickly assess the potential internal contamination in emergency situations. However, in the case that the analysis of multiple radionuclides is required in the same sample, time-consuming/tedious sequential analytical procedures using multiple chromatographic separation resins would have to be employed for the separation of every single radionuclide. In this work, a rapid method for the simultaneous determination of transuranium nuclides in urine was developed by using triple quadrupole inductively coupled plasma mass spectrometry (ICP-MS/MS) combined with a single DGA resin column. The chemical behaviors of Np/Pu and Am/Cm on the DGA resin were consistent in 8-10 mol/L HNO3 and 0.005-0.02 mol/L NaNO2 when 242Pu and 243Am were selected as tracers for Np/Pu and Am/Cm yield monitoring. Based on their different reaction rates with O2, 237Np, 239,240,241Pu, 241Am, and 244Cm in the same solution were simultaneously measured by ICP-MS/MS in the same run. The elimination efficiency of 238U+ tailing (7.43 × 10-9), 238U1H16O2+/238U16O2+ (8.11 × 10-8) and cross contamination of 241Pu and 241Am (<1%) were achieved using 10.0 mL/min He-0.3 mL/min O2 even if the eluate was directly measured without any evaporation. The detection limits of transuranium nuclides were at the femtogram level, demonstrating the feasibility of ICP-MS/MS for simultaneous transuranic radionuclides urinalysis. The developed method was validated by analyzing the spiked urine samples.

Iron quantification at the sub femtogram level in magnetite hybrid silica methacrylate core–shell nanocomposite particles by sp-ICP-MS

This study highlights the application of single particle (sp)-ICP-MS as a valuable tool to monitor the distribution ofiron oxide nanoparticles in hybrid organic–inorganic core-shell silica/PMMA nanocomposites.

Innovative Biosensing Approaches for Swift Identification of Candida Species, Intrusive Pathogenic Organisms.

Candida is the largest genus of medically significant fungi. Although most of its members are commensals, residing harmlessly in human bodies, some are opportunistic and dangerously invasive. These have the ability to cause severe nosocomial candidiasis and candidemia that affect the viscera and bloodstream. A prompt diagnosis will lead to a successful treatment modality. The smart solution of biosensing technologies for rapid and precise detection of Candida species has made remarkable progress. The development of point-of-care (POC) biosensor devices involves sensor precision down to pico-/femtogram level, cost-effectiveness, portability, rapidity, and user-friendliness. However, futuristic diagnostics will depend on exploiting technologies such as multiplexing for high-throughput screening, CRISPR, artificial intelligence (AI), neural networks, the Internet of Things (IoT), and cloud computing of medical databases. This review gives an insight into different biosensor technologies designed for the detection of medically significant Candida species, especially Candida albicans and C. auris, and their applications in the medical setting.

Metamaterial graphene sensors for the detection of two food additives.

Food safety is an important consideration for the food industry and for daily life, and food additives are essential in the modern food industry. Graphene-based metamaterial sensors are of great value and have potential applications in the detection of food additives, due to their ultra-sensitivity. This paper proposes a metasurface sensor consisting of graphene and dual elliptical ring resonators (Gr-DERRs) sensor for the detection of two common food additives. The limit of detection (LOD) for Sudan I solution is 581.43 fg/ml and, for taurine, 52.86 fg/ml. This ultra-sensitive detection is achieved by exploiting the unique electromagnetic properties of electromagnetically induced transparency (EIT) resonance, together with the Fermi energy level of graphene moving to the Dirac point, resulting in a dramatic change in the dielectric environment. The Gr-DERRs sensor has brings significant improvement in the detection of food additives with detection limits reduced to the femtogram level.

Simultaneous determination of femtogram levels of 237Np, 239Pu, and 240Pu in environmental solid samples using extraction chromatography and ICP-MS/MS

An analytical method based on triple quadrupole inductively coupled plasma mass spectrometry (ICP-MS/MS) was developed for simultaneous determination of femtogram levels of 237Np, 239Pu, and 240Pu in environmental samples. By carefully controlling the valence states of Np and Pu in the entire separation procedure using a simple single extraction chromatographic column (TK200), the consistent behavior of Np and Pu was achieved, allowing for the reliable application of 242Pu as the chemical yield tracer for 237Np, 239Pu, and 240Pu. A high decontamination factor of 3.2 × 107 for the most critical interfering element, uranium, was achieved in the chemical separation step. The interferences of 238U1H+ and peak tailing of 238U+ during the measurement of plutonium isotopes were effectively eliminated by utilizing 7.5 mL/min He–1.1 mL/min CO2 as reaction gases in the octupole collision/reaction cell and employing sequential quadrupole mode for mass separation in ICP-MS/MS. Specifically, the interference of 238U1H+ was reduced to 10−6 and the peak tailing of 238U+ to 10−10, surpassing the performance of measurement method without reaction gases by 3 orders of magnitude. The developed method enables the accurate determination of femtogram levels of 237Np, 239Pu, and 240Pu in the samples with U/Np and U/Pu atom ratios of up to 1017 and 1012, respectively. The developed method was validated by analyzing standard reference materials and spiked soil samples.

Evaluation of aptamer and molecularly imprinted polymers as a first hybrid sensor for leptin detection at femtogram levels

Selective and sensitive determination of macromolecules maintains its importance in diagnosing and determining diseases to protect human health. In this study, a hybrid sensor designed with dual recognition elements consisting of both aptamers (Apt) and molecularly imprinted polymers (MIPs) was carried out for the ultra-sensitive determination of Leptin. Firstly, the screen-printed electrode (SPE) surface was coated with platinum nanospheres (Pt NSs) and gold nanoparticles (Au NPs) to provide immobilization of the Apt[Leptin] complex on the surface. In the next step, the formed polymer layer around the complex using the electropolymerization of orthophenilendiamine (oPD) kept the Apt molecules on the surface more effectively. As expected, a synergistic effect occurred between the formed MIP cavities by removing Leptin from the surface and the embedded Apt molecules to fabricate a hybrid sensor. Under optimal conditions, responses in differential pulse voltammetry (DPV) currents showed a linear response over a wide concentration range from 1.0 fg/mL to 10.0 pg/mL with a limit of detection (LOD) of 0.31 fg/mL for Leptin detection. Moreover, the effectiveness of the hybrid sensor was assessed using real samples, such as human serum and plasma samples, and satisfactory recovery findings (106.2–109.0%) were found.

Simultaneous Determination of Transuranium Radionuclides for Nuclear Forensics by Compact Accelerator Mass Spectrometry.

Accelerator mass spectrometry (AMS) is one of the most sensitive techniques used to measure the long-lived actinides. This is particularly of interest for determination of ultra-trace transuranium nuclides and their isotopic fingerprints for nuclear forensics. In this work, a new method was developed for simultaneous determination of transuranium nuclides (Np, Pu, Am, and Cm isotopes) by using 300 kV AMS after a sequential chemical separation of each group of actinides. 242Pu and 243Am were utilized as tracers for Np/Pu and Am/Cm yield monitoring. The results show that the chemical behaviors of Np and Pu on the TK200 column and those of Am and Cm on the DGA column were very consistent in 8-9 mol/L of HNO3 and 0.015-0.03 mol/L of NaNO2 media during the radiochemical separation. The AMS detection efficiencies for transuranium nuclides were also evaluated. The detection limits for all radionuclides are below femtogram level and even in attogram level for Pu and Cm isotopes. The established method has been successfully applied to accurately measure various transuranium nuclides in a single actinide radionuclide solution, demonstrating its feasibility for nuclear forensic investigation.

Dynamic analysis of boron nitride nanotube using different boundary conditions under influence of vacancy defect: Insights from finite element method

In this paper, the effect of defects such as atomic vacancies (single atom vacancies like boron atom(Vb) or nitrogen atom vacancy(Vn) and di-atomic vacancies(VB-N) corresponds to vacancy of one boron and adjacent one nitrogen atom) on the vibrational behaviour of a single-walled boron nitride nanotube (SWBNNT) are investigated using finite element modelling in context of their applicability as mass sensors. The change in resonance frequency produced by vacancy defects at various points throughout the span when a mass is connected to the tip has also been examined. Space frames with three-dimensional components and point masses have been used to explain the cantilevered armchair (5, 5) and zigzag (10, 0) SW-BNNT with changing vacancy defects. The position of atomic vacancy has been varied (i.e., at 25 %, 50 % and 90 % of length from fixed end) to investigate the effect of location of defect along the length on the resonance behaviour of SW-BNNT. Further defective BNNTs have been analysed in such a way that the same can be used as nanomechanical resonator for detecting small mass of the order of femtogram level. For this,a small mass (varying from 10-20 to 10−25kg) is attached at the tip of cantilevered SW-BNNT. Substantial drop in frequency shift is observed when the position of defect moves toward the free end. the excitation frequency of defective BNNT is larger near the free end as compared to pristine one.

Trends in environmental monitoring of high explosives present in soil/sediment/groundwater using LC-MS/MS.

Environmental contamination by explosives occurs due to improper handling and disposal procedures. Explosives and their transformation products pose threat to human health and the ecosystem. Trace level detection of explosives present in different environmental matrices is a challenge, due to the interference caused by matrix components and the presence of cocontaminants. Liquid chromatography combined with tandem mass spectrometry (LC-MS/MS) is an advanced analytical tool, which is ideal for quantitative and qualitative detection of explosives and its metabolites at trace levels. This review aims to showcase the current trends in the application of LC-MS/MS for detecting explosives present in soil, sediment, and groundwater with detection limits ranging from nano to femtogram levels. Specificity and advantages of using LC-MS/MS over conventional analytical methods and various processing methods and techniques used for sample preparation are discussed in this article. Important application aspects of LC-MS/MS on environmental monitoring include site characterization and degradation evaluation. Studies on qualitative and quantitative LC-MS/MS analysis in determining the efficiency of treatment processes and contamination mapping, optimized conditions of LC and MS/MS adopted, role of different ionization techniques and mass analyzers in detection of explosives and its metabolites, relative abundance of various product ions formed on dissociation and the levels of detection achieved are reviewed. Ionization suppression, matrix effect, additive selection are some of the major factors which influence MS/MS detection. A summary of challenges and future research insights for effective utilization of this technique in the environmental monitoring of explosives are presented.

Automated Paper-Based Femtogram Sensing Device for Competitive Enzyme-Linked Immunosorbent Assay of Aflatoxin B1 Using Submicroliter Samples.

Microfluidic paper-based analytical devices (μPADs) are promising biosensors that may be used in a variety of bioanalytical applications. A μPAD for automating the competitive enzyme-linked immunosorbent assay (ELISA) of small-sized target detection at the femtogram level using submicroliter samples is reported in this study. The proposed μPAD was integrated with a sucrose valve to automate the sequential delivery of reagents, providing simple control of reagent delivery time and simple operation. The use of a sample solution dropping location at the zones on the device that had been prepared with an antibody-conjugated enzyme before immersion in a running buffer allowed minimization of sample volume to 0.6 μL, while eliminating the possible loss of a target molecule by adsorption on the membrane, thus improving detection sensitivity. Furthermore, the proposed device was successfully applied to the automation of competitive ELISA for the detection of aflatoxin B1 (AFB1), a potent carcinogen that causes substantial health risks to humans worldwide, with a detection limit of 60 femtograms or 0.1 ng/mL. The method developed in this study provides high sensitivity, small sample volume, on-site and equipment-free measurements, low-cost operation, and user-friendliness. This approach could be used to analyze small-sized molecules in the fields of food safety and quality control, environmental monitoring, and clinical diagnostics.

(Invited) Sensors for Agricultural Pests Management

Pests are undesired living organisms which cause major threat to plants, animals, humans and other organisms. It can directly feed on the crops and indirectly spread diseases and decrease the productivity of plants. Traditionally we have been using pesticides to get rid of them, but these pesticides are chemicals which tend to accumulate in the soil and water and pose detrimental effects to all the living creatures. To reduce the major loss in food production and use of chemical pesticides, we have come up with a few innovative approaches to control pests and also detect the toxic pesticides in an easy, affordable and eco-friendly way using sensors. We had developed sensors for effective use of biopesticides such as Helicoverpa armigera nuclear polyhedrosis virus (HaNPV), Spodoptera litura nuclear polyhedrosis virus (SlNPV) and Spilosoma obliqua nuclear polyhedrosis virus (SpilNPV). We had also designed a silicon dioxide-based microelectro-mechanical system (MEMS) sensors as tool for an early selective pest detection of female sex pheromones of Helicoverpa armigera and Bactrocera oleae. This robust device can also be used to selectively measure the pheromone at femtogram levels, and it is handy for early detection of pests. The device works well in both rainy and peak summer seasons. Using these systems farmers can take necessary measures in the pest attack region without using pesticides and would be able to support organic farming.

Detection of Microbial Agents in Oropharyngeal and Nasopharyngeal Samples of SARS-CoV-2 Patients.

The novel coronavirus outbreak started in December 2019 and rapidly spread around the globe, leading to a global pandemic. Here we reported the association of microbial agents identified in oropharyngeal and nasopharyngeal samples from patients with SARS-CoV-2 infection, using a Pan-microarray based technology referred to as PathoChIP. To validate the efficiency of PathoChIP, reference viral genomes obtained from BEI resource and 25 SARS-CoV-2 positive clinical samples were tested. This technology successfully detected femtogram levels of SARS-CoV-2 viral RNA, which demonstrated greater sensitivity and specificity than conventional diagnostic techniques. Simultaneously, a broad range of other microorganisms, including other viruses, bacteria, fungi and parasites can be detected in those samples. We identified 7 viral, 12 bacterial and 6 fungal agents common across all clinical samples suggesting an associated microbial signature in individuals who are infected with SARS-CoV-2. This technology is robust and has a flexible detection methodology that can be employed to detect the presence of all human respiratory pathogens in different sample preparations with precision. It will be important for differentiating the causative agents of respiratory illnesses, including SARS-CoV-2.

Hybrid approach for dynamic analysis of single-walled Boron Nitride Nanotube (SW-BNNT) in presence of vacancy defect

The appropriateness of single walled Boron Nitride Nanotube for recognition of mass as little as of the request for femtogram level (10–18 fg) in nearness of centre found point defect has been investigated through current paper using 3-dimensional atomistic structure approach based on finite element method. The impact of shifting mass in nearness of defect on frequency for armchair (5,5) Boron Nitride Nanotube has been analysed for bridge setup.Considering FEM approach the mass affectability for bridge design is seen as higher when contrasted with cantilever one. The experimental results delight that shorter length of BNNT with bridge arrangement is appropriate for detecting mass as little as of the request for femtogram level when mass is available on focus.

Urinary PCA3 detection in prostate cancer by magnetic nanoparticles coupled with colorimetric enzyme-linked oligonucleotide assay.

PCA3 is one of the most prostate cancer-specific genes described to date. Of note, PCA3 expression is detectable at high level in the urine of prostate cancer (PCa) patients. Accordingly, PCA3 is an ideal biomarker for PCa diagnosis. Several techniques for the measurement of this biomarker in urine have been developed but there are still some drawbacks. In this study, magnetic nanoparticle-based PCR coupled with streptavidin-horseradish peroxidase and a substrate for colorimetric detection was established as a potential assay for urinary PCA3 detection. The method provided a high specificity for PCA3 gene in LNCaP prostate cancer cell line. Additionally, this technique could detect PCA3 at femtogram level which was approximately 1,000-fold more sensitive than the conventional RT-PCR followed by agarose gel electrophoresis. The effectiveness of the method was assessed by PCA3 detection in clinical specimens. The relative PCA3 expression of PCa patients determined by this assay was significantly greater than that of benign prostatic hyperplasia (BPH) patients and healthy controls. The results of our test were comparable with the results of qRT-PCR. The proposed method is promising to distinguish between cancerous and non-cancerous groups. Altogether, this simple assay is practicable and useful for prostate cancer diagnosis.

Development of a Rapid and Sensitive IDH1/2 Mutation Detection Method for Glial Tumors and a Comparative Mutation Analysis of 236 Glial Tumor Samples.

The presence of mutations in the isocitrate dehydrogenase 1 and 2 genes (IDH1/2) in glioma tumors is correlated with good prognosis upon standard-of-care treatment. Therefore, information on whether the glioma tumor has IDH1/2 mutations could be used in the correct diagnosis and management of glial tumors. The two most common techniques used to detect IDH1/2 mutations, immunohistochemistry (IHC) and Sanger sequencing, are prone to missing these mutations, especially if the tumor cells that carry the mutations constitute a small minority of the tumor itself. We developed and validated a rapid method (3-mismatch-amplification refractory mutation system [3m-ARMS]) that can be used for pre-, intra- and postoperative detection of the most common IDH1/2 mutations in glial tumors with high specificity and sensitivity. We also conducted a comprehensive IDH1/2 mutation analysis in 236 glial tumor samples comparing 3m-ARMS, IHC and Sanger sequencing. 3m-ARMS was optimized and validated for the specific and sensitive detection of the most common IDH1 and IDH2 mutations. We then analyzed 236 glial tumor samples for the presence of IDH1/2 mutations using 3m-ARMS, Sanger sequencing and IHC techniques. We then analyzed and compared the results, evaluating the diagnostic and screening potential of 3m-ARMS. Comparison of the three techniques used in the mutation analysis showed that 3m-ARMS-based IDH1/2 mutation detection was superior to IHC and Sanger sequencing-based IDH1/2 mutation detection in terms of accuracy, specificity and sensitivity, especially for tumor samples in which only a small minority of the cell population carried the mutation. 3m-ARMS could detect the presence of femtogram levels of IDH1/2 mutant DNA in DNA samples in which the mutant DNA-to-wild-type DNA ratio was as low as 1:100,000. Sanger sequencing and IHC-based methods have shortcomings when detecting mutations in glial tumors so can miss IDH1/2 mutations in glial tumors when used alone without proper modifications. 3m-ARMS-based mutation detection is fast and simple with potential for use as a diagnostic test for the majority of hot spot mutations in IDH1/2 genes. It can detect IDH1/2 mutations within an hour so can be adapted for intraoperative diagnosis.

Ultra-sensitive detection of parathyroid hormone in human serum: a cheap and practical biosensing platform modified by an epoxy ended-silane agent

ABSTRACTIn this study, we designed an impedimetric immunosensor by modification of disposable ITO-PET (indium tin oxide-polyethylene terephthalate) electrode surface with 3-glycidoxypropyltriethoxysilane (3-GOPE). 3-GOPE silanization agent generates a suitable platform on the ITO-PET working electrode surface for antibody immobilisation and allows a wide determination range for analyte concentrations with the fabricated biosensor. All immobilisation processes, investigation of optimum parameters and characterisation studies of the developed immunosensor were evaluated and followed by Electrochemical Impedance Spectroscopy (EIS) and Cyclic Voltammetry (CV). Furthermore, the interactions between anti-parathyroid hormone (anti-PTH) and Parathyroid hormone (PTH) were simultaneously analysed with the Single Frequency Impedance (SFI) technique. All parameters which affect the biosensor response were optimised. A great wide concentration range (1 fg mL−1-2500 fg mL−1) and ultra-low detection limit (0.89 fg mL−1) were obtained under optimum conditions. Moreover, the impedimetric biosensor allowed high sensitivity in the detection of PTH in human serum. It was proved that the fabricated biosensor system has some advantages such as long-term storage life, very low limit of detection, ability to detect the PTH concentrations at femtogram level (1 fg mL−1- 2500 fg mL−1), excellent reproducibility and repeatability, reusability and ultra-high sensitivity.

Quantitative Imaging of Silver Nanoparticles and Essential Elements in Thin Sections of Fibroblast Multicellular Spheroids by High Resolution Laser Ablation Inductively Coupled Plasma Time-of-Flight Mass Spectrometry.

We applied high resolution laser ablation inductively coupled plasma time-of-flight mass spectrometry (LA-ICP-TOF-MS) with cellular spatial resolution for bioimaging of nanoparticles uptaken by fibroblast multicellular spheroids (MCS). This was used to quantitatively investigate interactions of silver nanoparticles (Ag NPs) and the distributions of intrinsic minerals and biologically relevant elements within thin sections of a fibroblast MCS as a three-dimensional in vitro tissue model. We designed matrix-matched calibration standards for this purpose and printed them using a noncontact piezo-driven array spotter with a Ag NP suspension and multielement standards. The limits of detection for Ag, Mg, P, K, Mn, Fe, Co, Cu, and Zn were at the femtogram (10-15 g) level, which is sufficient to investigate intrinsic minerals in thin MCS sections (20 μm thick). After incubation for 48 h, Ag NPs were enriched in the outer rim of the MCS but not detected in the core. The localization of Ag NPs was inhomogeneous in the outer rim, and they were colocalized with a single-cell-like structure visualized by Fe distribution (pixel size of elemental images: 5 × 0.5 μm). The quantitative value for the total mass of Ag NPs in a thin section by the present method agreed with that obtained by ICP-sector field (SF)-MS with a liquid mode after acid digestion.

Rapid concentration and isotopic measurements of ultra-trace 235U fission products with comparison to an ORIGEN isotope depletion model

This article presents the application of an automated online separation–direct analysis method, RAPID (Rapid Analysis of Post-Irradiation Debris), for the simultaneous measurement of both radioactive and stable fission isotopes from an irradiated highly-enriched uranium target. Developed for the measurement of the concentration and isotopic composition of over 40 elements down to the femtogram level, the RAPID method possesses the sensitivity, stability, and precision required to achieve accurate, low-level analyses of elements of non-natural origin. The isotopic compositions and concentrations of key fission elements cesium, strontium, yttrium, lanthanum, cerium, praseodymium, neodymium, promethium, and samarium have been measured repeatedly over a six-week period. The validity of these measurements was confirmed using ORIGEN (Oak Ridge Isotope GENeration), an isotope depletion and decay modeling software, to within 1–3%.