scholarly journals Development of a Novel, Rapid IntegratedCryptosporidium parvum Detection Assay

2000 ◽  
Vol 66 (7) ◽  
pp. 2711-2717 ◽  
Author(s):  
Diane Kozwich ◽  
Kristine A. Johansen ◽  
Keli Landau ◽  
Christopher A. Roehl ◽  
Sam Woronoff ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Solid Phase ◽  
Detection System ◽  
Immunomagnetic Separation ◽  
Immunofluorescence Assay ◽  
Lateral Flow ◽  
Reverse Transcription Pcr ◽  
Extraction And Purification ◽  
Flow Detection ◽  
Solid Phase Material

ABSTRACT The aim of this study was to develop a reverse transcription-PCR assay and lateral flow detection protocol for specific identification of Cryptosporidium parvum. The method which we developed is sensitive and specific and has a low limit of detection. In our protocol a solid phase material, the Xtra Bind Capture System, was used for extraction and purification of double-stranded RNA (dsRNA) specific for C. parvum. The Xtra Bind Capture System interfaced with pellets concentrated from water samples collected with previously developed filtration devices. The pellets were resuspended in reagent water (final volume, 0.5 ml), and an equal amount of rupture buffer and the Xtra Bind Capture System was added to the resuspended pellet mixture. The dsRNA target sequences in a 0.5-ml portion were captured by the solid phase material via hybridization. The debris and potential inhibitors were removed by washing the Xtra Bind material several times with buffer. The Xtra Bind material with its bound dsRNA was added directly to an amplification reaction mixture, and the target was amplified without elution from the Xtra Bind material. A PCR was performed in the presence of the Xtra Bind Capture System, which resulted in robust amplification of the target. The detection system which we used was adapted from lateral flow chromatography methods typically used for antigen-antibody reactions. The result was a colored line that was visible if the organism was present. When this method was used, we were able to reproducibly and correctly identify 10 oocysts added to 0.5 ml of reagent water. When the protocol was evaluated with a small set of environmental samples, the level of detection was as low as 1 oocyst/liter. The total time from resuspension of the pellet to detection was about 3 h, which is considerably less than the 5 h required for immunomagnetic separation followed by an indirect immunofluorescence assay and microscopy.

scholarly journals Carbon-Coated Superparamagnetic Nanoflowers for Biosensors Based on Lateral Flow Immunoassays

Biosensors ◽  
2020 ◽  
Vol 10 (8) ◽  
pp. 80
Author(s):  
Amanda Moyano ◽  
Esther Serrano-Pertierra ◽  
María Salvador ◽  
José Martínez-García ◽  
Yolanda Piñeiro ◽  
...  
Keyword(s):  
Magnetic Properties ◽  
Limit Of Detection ◽  
Detection System ◽  
Rapid Test ◽  
Superparamagnetic Iron Oxide ◽  
Extracellular Vesicle ◽  
Lateral Flow ◽  
Lateral Flow Immunoassay ◽  
Clear Correlation ◽  
One Pot

Superparamagnetic iron oxide nanoflowers coated by a black carbon layer (Fe3O4@C) were studied as labels in lateral flow immunoassays. They were synthesized by a one-pot solvothermal route, and they were characterized (size, morphology, chemical composition, and magnetic properties). They consist of several superparamagnetic cores embedded in a carbon coating holding carboxylic groups adequate for bioconjugation. Their multi-core structure is especially efficient for magnetic separation while keeping suitable magnetic properties and appropriate size for immunoassay reporters. Their functionality was tested with a model system based on the biotin–neutravidin interaction. For this, the nanoparticles were conjugated to neutravidin using the carbodiimide chemistry, and the lateral flow immunoassay was carried out with a biotin test line. Quantification was achieved with both an inductive magnetic sensor and a reflectance reader. In order to further investigate the quantifying capacity of the Fe3O4@C nanoflowers, the magnetic lateral flow immunoassay was tested as a detection system for extracellular vesicles (EVs), a novel source of biomarkers with interest for liquid biopsy. A clear correlation between the extracellular vesicle concentration and the signal proved the potential of the nanoflowers as quantifying labels. The limit of detection in a rapid test for EVs was lower than the values reported before for other magnetic nanoparticle labels in the working range 0–3 × 107 EVs/μL. The method showed a reproducibility (RSD) of 3% (n = 3). The lateral flow immunoassay (LFIA) rapid test developed in this work yielded to satisfactory results for EVs quantification by using a precipitation kit and also directly in plasma samples. Besides, these Fe3O4@C nanoparticles are easy to concentrate by means of a magnet, and this feature makes them promising candidates to further reduce the limit of detection.

Molecularly imprinted polymers for the selective determination of trace bisphenol A in river water by electrochemiluminescence

2013 ◽  
Vol 91 (8) ◽  
pp. 656-661 ◽  
Author(s):  
Xiaohua Jiang ◽  
Wengjie Ding ◽  
Chonglin Luan
Keyword(s):  
Bisphenol A ◽  
River Water ◽  
Limit Of Detection ◽  
Solid Phase ◽  
High Sensitivity ◽  
Selective Extraction ◽  
Molecularly Imprinted ◽  
Selective Determination ◽  
Extraction And Purification

The detection of bisphenol A (BPA) is very important for public health and environmental monitoring. In this work, BPA was found to be able to significantly quench the electrochemiluminescence (ECL) of the Ru(bpy)32+/2-(dibutylamino)ethanol (DBAE) system. Molecularly imprinted polymer was synthesized as solid-phase extraction sorbents, which were used for the selective extraction and purification of BPA. Under optimal conditions, the inhibited ECL intensity versus the logarithm of the concentration of BPA was in good linear relationship over a concentration range from 2.2 × 10−10 to 1.1 × 10−7 mol/L. The limit of detection was 4.5 × 10−11 mol/L (S/N = 3). The developed method was successfully applied for determination of BPA in river water with high sensitivity and reliability. Further, a possible mechanism for the quenching effects of the Ru(bpy)32+/DBAE system by BPA was also proposed.

Lateral-Flow Assay for Rapid Quantitative Detection of Clorprenaline Residue in Swine Urine

2014 ◽  
Vol 77 (10) ◽  
pp. 1824-1829 ◽  
Author(s):  
TAO PENG ◽  
FU S. ZHANG ◽  
WAN C. YANG ◽  
DONG X. LI ◽  
YUAN CHEN ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Detection System ◽  
Cold Treatment ◽  
Lateral Flow ◽  
Urine Samples ◽  
Feed Additive ◽  
Lateral Flow Immunoassay ◽  
Quantitative Detection ◽  
Swine Urine ◽  
Liquid Chromatography Tandem Mass

Clorprenaline (CLP), a β2-adrenergic agonist, was first found in veterinary drugs for cold treatment in China in 2013. It is a potential new lean meat-boosting feed additive because it can promote animal muscular mass growth and decrease fat accumulation. A competitive colloidal gold-based lateral flow immunoassay system with a portable strip reader was successfully developed for rapid quantitative detection of CLP residue in swine urine. The detection system was optimized so that the detection can be completed within 9 min with a limit of detection of 0.15 μg·liter−1. The assay exhibited good linear range from 3.0 to 20.0 μg·liter−1, with reliable correlation of 0.9970 and with no obvious cross-reaction with five other β2-agonist compounds. Twenty spiked swine urine samples were tested by lateral flow immunoassay and liquid chromatography–tandem mass spectrometry to confirm the accuracy of the system. Results show good correlation between the two methods. This method is rapid, sensitive, specific, and convenient. It can be applied in the field for on-site detection of CLP in urine samples.

scholarly journals Magnetic Lateral Flow Immunoassays

Diagnostics ◽  
2020 ◽  
Vol 10 (5) ◽  
pp. 288 ◽  
Author(s):  
Amanda Moyano ◽  
Esther Serrano-Pertierra ◽  
María Salvador ◽  
José Carlos Martínez-García ◽  
Montserrat Rivas ◽  
...  
Keyword(s):  
Magnetic Nanoparticles ◽  
Limit Of Detection ◽  
Current Knowledge ◽  
Immunomagnetic Separation ◽  
Sensitivity Enhancement ◽  
Lateral Flow ◽  
Rapid Separation ◽  
Latex Beads ◽  
New Generation

A new generation of magnetic lateral flow immunoassays is emerging as powerful tool for diagnostics. They rely on the use of magnetic nanoparticles (MNP) as detecting label, replacing conventional gold or latex beads. MNPs can be sensed and quantified by means of external devices, allowing the development of immunochromatographic tests with a quantitative capability. Moreover, they have an added advantage because they can be used for immunomagnetic separation (IMS), with improvements in selectivity and sensitivity. In this paper, we have reviewed the current knowledge on magnetic-lateral flow immunoassay (LFIA), coupled with both research and commercially available instruments. The work in the literature has been classified in two categories: optical and magnetic sensing. We have analysed the type of magnetic nanoparticles used in each case, their size, coating, crystal structure and the functional groups for their conjugation with biomolecules. We have also taken into account the analytical characteristics and the type of transduction. Magnetic LFIA have been used for the determination of biomarkers, pathogens, toxins, allergens and drugs. Nanocomposites have been developed as alternative to MNP with the purpose of sensitivity enhancement. Moreover, IMS in combination with other detection principles could also improve sensitivity and limit of detection. The critical analysis in this review could have an impact for the future development of magnetic LFIA in fields requiring both rapid separation and quantification.

Skin exposure: requirements to methods determining active ingredient of pesticides in washings

2019 ◽  
pp. 43-48
Author(s):  
V. N. Rakitskii ◽  
N. E. Fedorova ◽  
I. V. Bereznyak ◽  
N. G. Zavolokina ◽  
L. P. Muhina
Keyword(s):  
Lower Limit ◽  
Limit Of Detection ◽  
Solid Phase ◽  
Absorbed Dose ◽  
Ambient Air ◽  
Safety Coefficient ◽  
Limit Value ◽  
Calculated Risk ◽  
Nature Studies ◽  
Active Substances

The article presents results of studies exemplified by diquat on analysis concerning influence of lower limit value of quantitative assessment in washing sample for safety coefficient in exposure and in absorbed dose, if acting substance is absent in workplace ambient air samples and in dermal washings of workers. To control diquat in dermal washings, there is a method based on ion-pair liquid chromatography with ultraviolet detection (working wavelength 310 nm). To concentrate sample, cartridges for solid-phase extraction, containing ion exchange sorbent (Oasis MCX 6cc/500 mg), are used. Lower limit of assessment in washing sample — 0,15 micrograms. Experimentally set washing completeness is within range of 80–92%, standard deviation of repetition is 7,0% at most. The method created was tested in nature studies determining dermal exposure in workers subjected to 5 various preparations based on diquat dibromide when used for surface spraying from tractor and from aircraft. For lower limit of detection in washing sample (0,15 micrograms/washing), calculated risk value of exposure varied within 0,26–0,36; risk of absorbed dose was low — 0,23 (the allowable one ≤1). Findings are that present measuring methods which provide lower limit of detection 1 and 5 micrograms in washing sample could result in unallowable risk establishment even with absence of the substance in all samples of workplace air and dermal washings. The calculation formula suggested enables to give theoretic basis for requirements to lower limit of detecting active substances in dermal washing samples for evaluating risk of pesticides use in agriculture.

A Point of Care Lateral Flow Assay for Rapid and Colorimetric Detection of Interleukin 6 and Perspectives in Bedside Diagnostics

2020 ◽  
Author(s):  
Carla Eiras
Keyword(s):  
Interleukin 6 ◽  
Limit Of Detection ◽  
Inflammatory Diseases ◽  
Point Of Care ◽  
Colorimetric Detection ◽  
Lateral Flow ◽  
Lateral Flow Assay ◽  
Aggregation Assay ◽  
Before And After ◽  
Bedside Diagnosis

Interleukin-6 (IL-6) is a multifunctional cytokine and high bloodstream levels of which have been associated with severe inflammatory diseases, such as dengue fever, sepsis, various cancers, and visceral leishmaniasis (VL). Rapid tests for the quantification of IL-6 would be of great assistance for the bedside diagnosis and treatment of diseases such as VL. We have developed a lateral flow assay (LFA) for rapid and colorimetric IL-6 detection, consisting of anti-IL-6 antibodies conjugated to gold nanoparticles (AuNPs). The optimal concentration of anti-IL-6 used in the conjugate was determined to be 800.0 μg/mL, based on an aggregation assay using LFA. A linear relationship between IL-6 standard concentration and color intensity was observed after 20 min, with a linear range between 1.25 ng/mL and 9,000 ng/mL. The limit of detection for this method was estimated a t0.38 ng/mL. The concentration of IL-6 in five patients with severe VL was measured using LFA, and the results were consistent with those obtained using the cytometric bead array (CBA) method. A thorough analysis of the LFA membranes’ surface morphology, before and after sample contact, was performed using atomic force microscopy (AFM).The prototype described here is still being tested and improved, but this LFA will undoubtedly be of great help in the clinical quantification of IL-6.

A Point of Care Lateral Flow Assay for Rapid and Colorimetric Detection of Interleukin 6 and Perspectives in Bedside Diagnostics

2020 ◽  
Author(s):  
Carla Eiras
Keyword(s):  
Interleukin 6 ◽  
Limit Of Detection ◽  
Inflammatory Diseases ◽  
Point Of Care ◽  
Colorimetric Detection ◽  
Lateral Flow ◽  
Lateral Flow Assay ◽  
Aggregation Assay ◽  
Before And After ◽  
Bedside Diagnosis

Interleukin-6 (IL-6) is a multifunctional cytokine and high bloodstream levels of which have been associated with severe inflammatory diseases, such as dengue fever, sepsis, various cancers, and visceral leishmaniasis (VL). Rapid tests for the quantification of IL-6 would be of great assistance for the bedside diagnosis and treatment of diseases such as VL. We have developed a lateral flow assay (LFA) for rapid and colorimetric IL-6 detection, consisting of anti-IL-6 antibodies conjugated to gold nanoparticles (AuNPs). The optimal concentration of anti-IL-6 used in the conjugate was determined to be 800.0 μg/mL, based on an aggregation assay using LFA. A linear relationship between IL-6 standard concentration and color intensity was observed after 20 min, with a linear range between 1.25 ng/mL and 9,000 ng/mL. The limit of detection for this method was estimated at a t0.38 ng/mL. The concentration of IL-6 in five patients with severe VL was measured using LFA, and the results were consistent with those obtained using the cytometric bead array (CBA) method. A thorough analysis of the LFA membranes’ surface morphology, before and after sample contact, was performed using atomic force microscopy (AFM). The prototype described here is still being tested and improved, but this LFA will undoubtedly be of great help in the clinical quantification of IL-6.

An optimised and standardised test to determine the presence of the protozoa Cryptosporidium and Giardia in water

2000 ◽  
Vol 41 (7) ◽  
pp. 103-110 ◽  
Author(s):  
G. Stanfield ◽  
E. Carrington ◽  
F. Albinet ◽  
B. Compagnon ◽  
N. Dumoutier ◽  
...  
Keyword(s):  
Solid Phase ◽  
Immunomagnetic Separation ◽  
Ferric Sulphate ◽  
Preliminary Assessment ◽  
New Methods ◽  
European Water ◽  
Conventional Analysis ◽  
Time Savings ◽  
Initial Recovery ◽  
Microscopic Counting

With funding from the European Commission, a consortium of members of the European Water Research Institutes is carrying out a programme of work with the objective of optimising and standardising a method for determining the presence in water of (oo)cysts of Cryptosporidium and Giardia. Each of the stages of the conventional analysis procedure (initial concentration, recoveryand identification and enumeration) are being investigated and the relative merits of existing and new methods are being assessed. Newly developed filters (Envirochek and Filta-Max) have been shown to be more efficient for initial recovery of (oo)cysts from water than the previously used Cuno cartridge filters. In addition, for the analysis of raw waters, flocculationwith ferric sulphate has been shown to give recoveries similar to the Envirochek and Filta Max. Modern purification systems such as immunomagnetic separation have also been assessed and found to offer some advantages over flotation although optimisation of the latter has brought improved efficiency. Preliminary assessment of solid phase cytometry has indicated that this technique could offer significant time savings compared to conventional microscopic counting. The results of the study will be used to propose a revised standard method to CEN.

Calcium/Copper alginate framework doped with CuO nano-particles as a novel adsorbent for micro-extraction of benzodiazepines from human serum

2020 ◽  
Vol 16 ◽  
Author(s):  
Nadereh Rahbar ◽  
Fatemeh Ahmadi ◽  
Zahra Ramezani ◽  
Masoumeh Nourani
Keyword(s):  
Human Serum ◽  
High Performance ◽  
Limit Of Detection ◽  
Solid Phase ◽  
Nano Particles ◽  
Limit Of Quantification ◽  
Analytical Methodology ◽  
Fiber Fabrication ◽  
Relative Standard ◽  
Green Procedure

Background: Sample preparation is one of the most challenging phases in pharmaceutical analysis, especially in biological matrices, affecting the whole analytical methodology. Objective: In this study, a new Ca(II)/Cu(II)/alginate/CuO nanoparticles hydrogel fiber (CCACHF) was synthesized through a simple, green procedure and applied for fiber micro solid phase extraction (FMSPE) of diazepam (DIZ) and oxazepam (OXZ) as model drugs prior to high-performance liquid chromatography-UV detection (HPLC-UV). Methods: Composition and morphology of the prepared fiber were characterized and the effect of main parameters on the fiber fabrication and extraction efficiency have been studied and optimized. Results: In optimal conditions, calibration curves were linear ranging between 0.1–500 µg L−1 with regression coefficients of 0.9938 and 0.9968. Limit of detection (LOD) (S/N=3) and limit of quantification (LOQ) (S/N=10) of the technique for DIZ and OXZ were 0.03 to 0.1 µg L−1. Within-day and between-day relative standard deviations (RSDs) for DIZ and OXZ were 6.0–12.5% and 3.3–9.4%, respectively. Conclusion: The fabricated adsorbent has been substantially employed to extraction of selected benzo-diazepines (BZDs) from human serum real specimens and the obtained recoveries were also satisfactory (82.1-109.7%).

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