scholarly journals Fluorescence Polarization Immunoassay for Determination of Enrofloxacin in Pork Liver and Chicken

Molecules ◽  
2019 ◽  
Vol 24 (24) ◽  
pp. 4462
Author(s):  
Xing Shen ◽  
Jiahong Chen ◽  
Shuwei Lv ◽  
Xiulan Sun ◽  
Boris B. Dzantiev ◽  
...  
Keyword(s):  
Fluorescence Polarization ◽  
Limit Of Detection ◽  
Polyclonal Antibodies ◽  
Screening Method ◽  
Sample Pretreatment ◽  
High Sensitivity ◽  
Cross Reactivity ◽  
Fluorescence Polarization Immunoassay ◽  
Pork Liver ◽  
Sensitivity Specificity

Enrofloxacin (ENR) is a widely used fluoroquinolone (FQ) antibiotic for antibacterial treatment of edible animal. In this study, a rapid and highly specific fluorescence polarization immunoassay (FPIA) was developed for monitoring ENR residues in animal foods. First, ENR was covalently coupled to bovine serum albumin (BSA) to produce specific polyclonal antibodies (pAbs). Three fluorescein-labeled ENR tracers (A, B, and C) with different spacers were synthesized and compared to obtain higher sensitivity. Tracer C with the longest arm showed the best sensitivity among the three tracers. The developed FPIA method showed an IC50 (50% inhibitory concentration) of 21.49 ng·mL−1 with a dynamic working range (IC20–IC80) of 4.30–107.46 ng·mL−1 and a limit of detection (LOD, IC10) of 1.68 ng·mL−1. The cross-reactivity (CR) of several structurally related compounds was less than 2%. The recoveries of spiked pork liver and chicken samples varied from 91.3% to 112.9%, and the average coefficients of variation were less than 3.83% and 5.13%, respectively. The immunoassay took only 8 min excluding sample pretreatment. This indicated that the established method had high sensitivity, specificity, and the advantages of simplicity. Therefore, the proposed FPIA provided a useful screening method for the rapid detection of ENR residues in pork liver and chicken.

scholarly journals Design, Synthesis, and Characterization of Tracers and Development of a Fluorescence Polarization Immunoassay for Rapid Screening of 4,4′-Dinitrocarbanilide in Chicken Muscle

Foods ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1822
Author(s):  
Qidi Zhang ◽  
Ming Zou ◽  
Wanyu Wang ◽  
Jinyan Li ◽  
Xiao Liang
Keyword(s):  
Fluorescence Polarization ◽  
Limit Of Detection ◽  
Screening Method ◽  
Sample Pretreatment ◽  
Cross Reactivity ◽  
Rapid Screening ◽  
Fluorescence Polarization Immunoassay ◽  
Design Synthesis ◽  
Chicken Muscle ◽  
Edible Tissues

The compound, 4,4′-dinitrocarbanilide (DNC), is the marker residue of concern in edible tissues of broilers fed with diets containing anticoccidial nicarbazin (NIC). In this study, 25 fluorescein-labeled DNC derivatives (tracers) are synthesized and characterized to develop a rapid fluorescence polarization immunoassay (FPIA) for the detection of DNC in chickens using DNC monoclonal antibodies (mAbs). The effect of the tracer structure on the sensitivity of the FPIA is investigated. Our results show that after optimization, the half maximal inhibitory concentrations (IC50) and limit of detection (LOD) of the FPIA in the buffer are 28.3 and 5.7 ng mL−1, respectively. No significant cross-reactivity (CR < 0.89%) with 15 DNC analogues is observed. The developed FPIA is validated for DNC detection in spiked chicken homogenates, and recoveries ranged from 74.2 to 85.8%, with coefficients of variation <8.6%. Moreover, the total time needed for the detection procedure of the FPIA, including sample pretreatment, is <40 min, which has not been achieved in any other immunoassays for DNC from literature. Our results demonstrate that the FPIA developed here is a simple, sensitive, specific, and reproducible screening method for DNC residues in chickens.

scholarly journals Fluorescence Polarization Immunoassay for the Determination of T-2 and HT-2 Toxins and Their Glucosides in Wheat

Toxins ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 380 ◽  
Author(s):  
Vincenzo Lippolis ◽  
Anna C. R. Porricelli ◽  
Erminia Mancini ◽  
Biancamaria Ciasca ◽  
Veronica M. T. Lattanzio ◽  
...  
Keyword(s):  
Fluorescence Polarization ◽  
Screening Method ◽  
High Sensitivity ◽  
Cross Reactivity ◽  
Screening Methods ◽  
The European Union ◽  
Fluorescence Polarization Immunoassay ◽  
Intermediate Precision ◽  
Relative Standard

T-2 and HT-2 toxins and their main modified forms (T-2 glucoside and HT-2 glucoside) may co-occur in cereals and cereal-based products. A fluorescence polarization immunoassay (FPIA) was developed for the simultaneous determination of T-2 toxin, HT-2 toxin and relevant glucosides, expressed as sum. The developed FPIA, using a HT-2-specific antibody, showed high sensitivity (IC50 = 2.0 ng/mL) and high cross-reactivity (100% for T-2 toxin and 80% for T-2 and HT-2 glucosides). The FPIA has been used to develop two rapid and easy-to-use methods using two different extraction protocols, based on the use of organic (methanol/water, 90:10, v/v) and non-organic (water) solvents, for the determination of these toxins in wheat. The two proposed methods showed analytical performances in terms of sensitivity (LOD 10 µg/kg) recovery (92–97%) and precision (relative standard deviations ≤13%), fulfilling the criteria for acceptability of an analytical method for the quantitative determination of T-2 and HT-2 toxins established by the European Union. Furthermore, the methods were then validated in accordance with the harmonized guidelines for the validation of screening methods included in the Regulation (EU) No. 519/2014. The satisfactory analytical performances, in terms of intermediate precision (≤25%), cut-off level (80 and 96 µg/kg for the two methods) and rate of false positives (<0.1%) confirmed the applicability of the proposed methods as screening method for assessing the content of these toxins in wheat at the EU indicative levels reported for T-2 and HT-2 toxins.

High Throughput Determination of BTEX by a One-Step Fluorescence Polarization Immunoassay

2005 ◽  
Vol 2 (3) ◽  
pp. 227 ◽  
Author(s):  
Sergei A. Eremin ◽  
Dietmar Knopp ◽  
Reinhard Niessner ◽  
Ji Youn Hong ◽  
Song-Ja Park ◽  
...  
Keyword(s):  
Fluorescence Polarization ◽  
Limit Of Detection ◽  
Petroleum Products ◽  
Cross Reactivity ◽  
Rapid Screening ◽  
Fluorescence Polarization Immunoassay ◽  
Phenylcarboxylic Acids ◽  
One Step ◽  
Benzene Toluene

Environmental Context.Benzene, toluene, ethylbenzene, and xylenes (BTEX) are used as solvents in paints and coatings and are constituents of petroleum products. BTEX can contaminate air, water or soil and is toxic; benzene, in particular, is a recognized human carcinogen. Most existing methods for detecting BTEX are time-consuming, complicated and very expensive for routine screening. A rapid immunoassay of BTEX is presented that greatly simplifies environmental monitoring of water contamination. Abstract.For the rapid screening of BTEX (benzene, toluene, ethylbenzene, xylenes), a fluorescence polarization immunoassay (FPIA) was developed using the fluorescence polarization analyzer Abbott TDx. Several fluorescence-labelled tracers were synthesized by binding ethylenediamine fluorescein thiocarbamyl (EDF) to various substituted phenylcarboxylic acids. The binding of 27 tracers with two antisera that can be considered as class-specific for BTEX was investigated to select optimal tracer–antibody pairs. Significant differences were found in binding, titer, sensitivity, and assay kinetics. A best pair of anti-BTEX antiserum and EDF-labelled p-tolylacetic acid tracer was selected for FPIA. To simplify the method, an immunocomplex single reagent was prepared to detect BTEX by a one-step FPIA. One-step FPIA is a rapid homogeneous type of immunoassay that has only one pipetting step, does not need separation of free and bound analyte and can be performed at room temperature. The within-run coefficient of variation was ranged between 3.4% and 5.7%. Furthermore, if the measurement can be done at constant temperature, standards for every assay run are unnecessary. Cross-reactivity studies of petroleum compounds and a BTEX mixture indicated that p-xylene was most reactive with a limit of detection (LOD) of 0.22 µg mL−1 in 50 µL of sample. The LOD for toluene and benzene was 2.1 and 11 µg mL−1 respectively. The immunocomplex single reagent has proven to be significantly more stable than the corresponding solutions of antibody and tracer.

Evaluation of an alternative S100b assay for use in cardiac surgery: relationship with microemboli and neuropsychological outcome

Perfusion ◽  
2007 ◽  
Vol 22 (4) ◽  
pp. 267-272 ◽  
Author(s):  
D.C. Whitaker ◽  
A.J.E. Green ◽  
J. Stygall ◽  
M.J.G. Harrison ◽  
S.P. Newman
Keyword(s):  
Cardiac Surgery ◽  
Limit Of Detection ◽  
Artery Bypass ◽  
Sandwich Elisa ◽  
High Sensitivity ◽  
Cross Reactivity ◽  
Cell Protein ◽  
Skin Closure ◽  
Neuropsychological Outcome ◽  
The Right

Introduction. The aim of the study was to investigate the relationship between S100b release, neuropsychological outcome and cerebral microemboli. Peri-operative assay of the astroglial cell protein S100b has been used as a marker of cerebral damage after cardiac surgery but potential assay cross-reactivity has limited its specificity. The present study uses an alternative enzyme-linked immunoabsorbant assay (ELISA) for serum S100b that has documented sensitivity and specificity data in patients undergoing coronary artery bypass grafting (CABG). Methods. Fifty-five consecutive patients undergoing routine CABG surgery received serial venous S100b sampling at five time points: i) Pre-operative, ii) At the end of cardiopulmonary bypass (CPB), iii) 6 hrs, iv) 24 hrs and v) 48 hrs post skin closure. A previously described sandwich ELISA with monoclonal anti- S100b was used. This assay has a lower limit of detection of 0.04 μ g/L and < 0.006% reactivity with S100a at a concentration of 100 μg/L S100a. Cerebral microemboli during surgery were recorded by transcranial Doppler monitor over the right middle cerebral artery. Evidence of cerebral impairment was obtained by comparing patients' performance in a neuropsychological battery of 9 tests administered 6—8 weeks post-operatively with their pre-operative scores. Results. There was a significant increase in S100b only at the end of bypass (mean 0.30 μg/L, SD ± 0.33 and range .00 to 1.57). S100b levels at the end of bypass did not correlate with neuropsychological outcome or microemboli counts. Conclusions. The low levels of S100b detected using the present assay, despite its high sensitivity and despite the routine use of cardiotomy suction, suggest that the assay may have higher specificity for cerebral S100b than previously used assays. There was no evidence that this assay is related to neuropsychological change or cerebral microemboli in cardiac surgery. Perfusion (2007) 22, 267—272.

Detection of Zearalenone and Related Metabolites by Fluorescence Polarization Immunoassay

2004 ◽  
Vol 67 (5) ◽  
pp. 1039-1043 ◽  
Author(s):  
CHRIS M. MARAGOS ◽  
EUN-KYUNG KIM
Keyword(s):  
Small Molecules ◽  
Fluorescence Polarization ◽  
Cross Reactivity ◽  
Fluorescence Polarization Immunoassay ◽  
Rapid Extraction ◽  
Culture Material ◽  
Liquid Chromatographic Method ◽  
The World ◽  
Enzyme Label ◽  
Liquid Chromatographic

Zearalenone is an estrogenic mycotoxin commonly found in grains throughout the world. A number of instrument- and antibody-based methods including enzyme-linked immunosorbent assays (ELISAs) have been developed to detect zearalenone (ZEN) and related toxins in commodities and foods. Although convenient, the commercial ELISAs for small molecules such as ZEN require a washing step to separate bound and unbound enzyme label before detection. In fluorescence polarization immunoassays, separation of bound and unbound label is not required, a property that reduces the time needed to perform the assays. We developed a fluorescence polarization immunoassay for ZEN in maize. When combined with a rapid extraction technique, the assay could be used to detect as little as 0.11 μg of ZEN g−1 maize within 10 min. The assay showed cross-reactivity to the ZEN analogs zearalanone, α-zearalanol, α-zearalenol, β-zearalenol, and β-zearalanol of 195, 139, 102, 71, and 20%, respectively, relative to ZEN (100%). Recovery of ZEN from spiked maize over the range of 0.5 to 5 μg g−1 averaged 100.2% (n = 12). The fluorescence polarization immunoassay results were comparable to those obtained with a liquid chromatographic method for the analysis of 60 naturally contaminated maize samples and maize samples amended with culture material. The fluorescence polarization immunoassay provides a rapid method for screening of maize for ZEN.

scholarly journals Rapid immunoassays for detection of anabolic nortestosterone in dietary supplements

2013 ◽  
Vol 31 (No. 5) ◽  
pp. 514-519 ◽  
Author(s):  
B. Holubová ◽  
S. Göselová ◽  
L. Ševčíková ◽  
M. Vlach ◽  
M. Blažková ◽  
...  
Keyword(s):  
Dietary Supplements ◽  
Rapid Detection ◽  
Visual Detection ◽  
Limit Of Detection ◽  
Polyclonal Antibodies ◽  
Cross Reactivity ◽  
Rapid Analysis ◽  
Experimental Conditions ◽  
Immunochromatographic Strip ◽  
Strip Test

An enzyme immunoassay (ELISA) and an immunochromatographic strip were designed for a rapid detection of nortestosterone in dietary supplements. Two polyclonal antibodies and two types of nortestosterone-protein coating conjugates were tested to develop the most appropriate method. Under optimal experimental conditions, the most sensitive ELISA achieved the IC<sub>50 </sub>and the limit of detection values of 6.41 and 0.09 ng/ml, respectively. The assay specificity was tested measuring cross-reactivity of several steroids. The interference with the assay was negligible (&lt; 0.1%), except for cross-reactivity with another frequently abused steroid testosterone (23%). The optimised gold particle-based immunochromatographic strip provided in semi-quantitative test a visual detection limit of 1&nbsp;ng/ml. None of these methods showed the interference using a filtrate of the suspension of non-contaminated sample. After the validation for particular matrices, the ELISA and the strip test could be useful tools for a rapid analysis of nortestosterone in crude extracts of dietary supplements.

scholarly journals 1089. Analytical Performance of an Ultrasensitive Immunoassay for Detection of Clostridium difficile Toxins in Stool

2018 ◽  
Vol 5 (suppl_1) ◽  
pp. S326-S326
Author(s):  
Amelita Bartolome ◽  
Anna Almazan ◽  
Salina Abusali ◽  
Stanley Tam ◽  
Eric Lee ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Room Temperature ◽  
High Sensitivity ◽  
Turnaround Time ◽  
Neutralization Assay ◽  
Cross Reactivity ◽  
Analytical Performance ◽  
Freeze Thaw ◽  
Gastrointestinal Pathogens ◽  
Sensitivity Specificity

Abstract Background Clostridium difficile infection (CDI) is the main cause for nosocomial diarrhea. Currently available assays for the diagnosis of CDI show deficits in sensitivity, specificity, and/or turnaround time. The Singulex Clarity® C. diff toxins A/B assay, in development for the Singulex Clarity® system, was designed to provide an accurate and automated detection of C. difficile toxins A (TcdA) and B (TcdB) in stool. Here, the analytical performance of the assay is reported. Methods Limits of detection (LoD) for TcdA and TcdB in stool and buffer was determined, and a preliminary cutoff, as compared with cell cytotoxicity neutralization assay (CCNA), was established. Analytical reactivity against 38 toxigenic and nontoxigenic C. difficile strains of eight different toxinotypes was determined. Cross-reactivity against 53 other gastrointestinal pathogens and potential interference by 11 endogenous and exogenous substances were determined. Reproducibility was tested with triplicate samples (n = 85), and stability was evaluated in samples stored at room temperature, refrigerated, and frozen conditions, and subjected to three freeze-thaw cycles. Results The LoDs for TcdA and TcdB were 0.8 and 0.3 pg/mL in buffer, and 2.0 and 0.7 pg/mL in stool, respectively. Using a preliminary cutoff, the assay demonstrated 96.3% sensitivity and 96.1% specificity compared with CCNA. The Singulex Clarity® C. diff toxins A/B assay detected toxins from all tested strains and toxinotypes. No cross-reactivity or interference were detected. The repeatability was 99%, and samples for C. difficile toxin testing were stable up to 8 hours in room temperature, 1 week in 2–8°C, 6 months in −70°C, and up to three freeze–thaw cycles. Conclusion The Singulex Clarity C. diff toxins A/B assay (in development) can detect TcdA and TcdB at very low concentrations and it has high sensitivity and specificity compared with CCNA. The assay demonstrates reactivity to common C. difficile strains, does not show cross-reactivity to common gastrointestinal pathogens, is robust against common interferents, allows for toxin detection in both fresh and frozen stool samples and up to three freeze–thaw cycles, and provides results with high reproducibility. Disclosures A. Bartolome, Singulex, Inc.: Employee, Salary. A. Almazan, Singulex, Inc.: Employee, Salary. S. Abusali, Singulex, Inc.: Employee, Salary. S. Tam, Singulex, Inc.: Employee, Salary. E. Lee, Singulex, Inc.: Employee, Salary. A. Changavi, Singulex, Inc.: Employee, Salary. W. Trinh, Singulex, Inc.: Employee, Salary. K. Chau, Singulex, Inc.: Employee, Salary. J. Estis, Singulex, Inc.: Employee, Salary. B. Noland, Singulex, Inc.: Employee, Salary. J. Bishop, Singulex, Inc.: Employee, Salary.

Monitoring tricyclic antidepressant concentrations in serum by fluorescence polarization immunoassay compared with gas chromatography and HPLC

1994 ◽  
Vol 40 (6) ◽  
pp. 929-933 ◽  
Author(s):  
M L Rao ◽  
U Staberock ◽  
P Baumann ◽  
C Hiemke ◽  
A Deister ◽  
...  
Keyword(s):  
Gas Chromatography ◽  
Fluorescence Polarization ◽  
Tricyclic Antidepressants ◽  
Limit Of Detection ◽  
Correlation Coefficients ◽  
Tricyclic Antidepressant ◽  
Therapeutic Drug ◽  
Fluorescence Polarization Immunoassay

Abstract The fluorescence polarization immunoassay (FPIA) developed by Abbott to diagnose intoxication with tricyclic antidepressants was adapted for therapeutic drug monitoring and validated with chromatograpic methods to investigate its potential for this use. We compared serum concentrations of tricyclic antidepressants in vivo and in vitro obtained by FPIA with those by gas chromatography and HPLC. For amitriptyline, imipramine, clomipramine, and doxepin, the detection limit of the FPIA was 72, 71, 64, and 72 nmol/L (approximately 20 micrograms/L), respectively; that by gas chromatography was 18, 18, and 16 nmol/L (approximately 5 micrograms/L) for amitriptyline, imipramine and clomipramine, respectively; with HPLC the lower limit of detection for doxepin was 36 nmol/L (10 micrograms/L). The intra- and interassay CVs ranged from 3% to 6%. In patients being treated with amitriptyline, imipramine, clomipramine, and doxepin, at steady-state the correlation coefficients between FPIA and GC/HPLC results for split samples were 0.95, 0.92, 0.90 and 0.70, respectively. However, the slopes were close to unity only for amitriptyline and doxepin, being 0.6 for imipramine and 1.9 for clomipramine.

Fluorescence polarization immunoassay evaluated for screening for amphetamine and methamphetamine in urine.

1987 ◽  
Vol 33 (7) ◽  
pp. 1200-1202 ◽  
Author(s):  
Y H Caplan ◽  
B Levine ◽  
B Goldberger
Keyword(s):  
Fluorescence Polarization ◽  
Cross Reactivity ◽  
Fluorescence Polarization Immunoassay ◽  
Limit Of Quantification ◽  
Immunoassay Technique ◽  
Urine Specimens ◽  
Lower Cutoff

Abstract We studied the recently developed Abbott fluorescence polarization immunoassay (FPIA) for amphetamine and methamphetamine in urine and compared the results with those of the Syva enzyme-multiplied immunoassay technique (EMIT) and a gas-chromatographic assay. The FPIA method showed a limit of quantification of 0.3 mg/L, comparable with the lower cutoff of the EMIT assay. FPIA demonstrated greater specificity than the EMIT assay: phenylpropanolamine and ephedrine showed extremely limited cross reactivity with the FPIA antibody. Analysis of 249 urine specimens by all three methods clearly demonstrated the FPIA method to be acceptable for screening for amphetamine and methamphetamine in urine.

scholarly journals Development of PCR, LAMP and qPCR Assays for the Detection of Aflatoxigenic Strains of Aspergillus flavus and A. parasiticus in Hazelnut

Toxins ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 757
Author(s):  
Sara Franco Ortega ◽  
Ilenia Siciliano ◽  
Simona Prencipe ◽  
Maria Lodovica Gullino ◽  
Davide Spadaro
Keyword(s):  
Food Safety ◽  
Real Time ◽  
Aspergillus Flavus ◽  
Real Time Pcr ◽  
Limit Of Detection ◽  
Screening Method ◽  
Lamp Assay ◽  
High Sensitivity ◽  
Isothermal Amplification

Aspergillus flavus and A. parasiticus are two species able to produce aflatoxins in foodstuffs, and in particular in hazelnuts, at harvest and during postharvest phase. As not all the strains of these species are aflatoxin producers, it is necessary to develop techniques that can detect aflatoxigenic from not aflatoxigenic strains. Two assays, a LAMP (loop-mediated isothermal amplification) and a real time PCR with TaqMan® probe were designed and validated in terms of specificity, sensitivity, reproducibility, and repeatability. The capability of the strains to produce aflatoxins was measured in vitro and both assays showed to be specific for the aflatoxigenic strains of A. flavus and A. parasiticus. The limit of detection of the LAMP assay was 100–999 picograms of DNA, while the qPCR detected 160 femtograms of DNA in hazelnuts. Both techniques were validated using artificially inoculated hazelnuts and naturally infected hazelnuts. The qPCR was able to detect as few as eight cells of aflatoxigenic Aspergillus in naturally infected hazelnut. The combination of the LAMP assay, which can be performed in less than an hour, as screening method, with the high sensitivity of the qPCR, as confirmation assay, is able to detect aflatoxigenic strains already in field, helping to preserve the food safety of hazelnuts.

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