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Topical Collection "Biorecognition Assays for Mycotoxins"

A topical collection in Toxins (ISSN 2072-6651). This collection belongs to the section "Mycotoxins".

Editors

Collection Editor
Dr. Michelangelo Pascale

Institute of Sciences of Food Production (ISPA), National Research Council of Italy (CNR), via G. Amendola 122/O, 70126 Bari, Italy
Website | E-Mail
Interests: Development and validation of analytical methods for mycotoxins
Collection Editor
Dr. Maria C. DeRosa

Department of Chemistry and Institute of Biochemistry, Carleton University, 225 Steacie Building, 1125 Colonel By Drive, Ottawa, ON K1S 5B6, Canada
Website | E-Mail
Phone: 613-520-2600 x. 3844
Interests: Aptamer-based biosensors and other DNA nanotechnology

Topical Collection Information

Dear Colleagues,

Mycotoxins are toxic fungal metabolites that contaminate several crops worldwide and could represent a significant hazard to human and animal health. Maximum levels for the major mycotoxins have been established in several commodities worldwide. Therefore, a continuous monitoring of these natural contaminants in food and feed is essential to prevent the risk of exposure. Liquid chromatographic methods, coupled with ultraviolet, fluorescence, or mass spectrometry detectors are commonly used for the determination of a large number of mycotoxins. However they are expensive, time-consuming, and require sample clean-up and trained personnel. There is a growing need for rapid and high-throughput screening methods to be used in the food/feed chains and this is reflected in the continued growth of global markets for such assays. Screening methods, based on bio-sensing technologies, are fast, inexpensive, and require visual evaluation or simple instruments. Enzyme immunosorbent assays, lateral flow devices, strip tests, flow-through immunoassays, fluorescence polarization immunoassays based on antibodies, aptamers, peptides or bio-mimetic recognition elements, i.e., molecularly imprinted polymers, are some examples of biorecognition assays for mycotoxin detection. This Special Issue aims to highlight recent developments in this important field.

Dr. Michelangelo Pascale
Asso. Prof. Maria C. DeRosa
Collection Editors

Submission

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Keywords

  • Mycotoxin
  • Immunoassay
  • Biorecognition
  • Antibody
  • Aptamer
  • Biosensor
  • Biosensing Technique
  • Lab-on-a-chip

Published Papers (18 papers)

2017

Jump to: 2016, 2015

Open AccessArticle T-2 Toxin/HT-2 Toxin and Ochratoxin A ELISAs Development and In-House Validation in Food in Accordance with the Commission Regulation (EU) No 519/2014
Toxins 2017, 9(12), 388; doi:10.3390/toxins9120388
Received: 15 November 2017 / Revised: 25 November 2017 / Accepted: 29 November 2017 / Published: 30 November 2017
PDF Full-text (1219 KB) | HTML Full-text | XML Full-text
Abstract
T-2 toxin/HT-2 toxin (T-2/HT-2) and ochratoxin A (OTA) are mycotoxins that can contaminate a variety of agricultural commodities. To protect consumers’ health, indicative limits for T-2/HT-2 and maximum limits for OTA have been set by the European Commission, requiring food business operators and
[...] Read more.
T-2 toxin/HT-2 toxin (T-2/HT-2) and ochratoxin A (OTA) are mycotoxins that can contaminate a variety of agricultural commodities. To protect consumers’ health, indicative limits for T-2/HT-2 and maximum limits for OTA have been set by the European Commission, requiring food business operators and controlling agencies to conduct routine checks for the presence of these harmful contaminants. Screening methods are increasingly used for monitoring purposes. Due to the demand for new and improved screening tools, two individual detection methods, T-2/HT-2 and OTA enzyme-linked immunosorbent assays (ELISAs), were developed in this study. The T-2/HT-2 ELISA was based on a T-2 monoclonal antibody with an IC50 (50% inhibitory concentration) of 0.28 ng/mL and 125% cross-reactivity with HT-2. As regards the OTA ELISA, a new sensitive monoclonal antibody specific to OTA with an IC50 of 0.13 ng/mL was produced. Both developed ELISA tests were then validated in agricultural commodities in accordance with the new performance criteria guidelines for the validation of screening methods for mycotoxins included in Commission Regulation (EU) No 519/2014. The T-2/HT-2 ELISA was demonstrated to be suitable for the detection of T-2/HT-2 in cereals and baby food at and above the screening target concentration (STC) of 12.5 μg/kg and 7.5 μg/kg, respectively. The OTA ELISA was shown to be applicable for the detection of OTA in cereals, coffee, cocoa and wine at and above the STC of 2 μg/kg, 2.5 μg/kg, 2.5 μg/kg and 0.4 ng/mL, respectively. The accuracy of both ELISAs was further confirmed by analysing proficiency test and reference samples. The developed methods can be used for sensitive and high-throughput screening for the presence of T-2/HT-2 and OTA in agricultural commodities. Full article
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Open AccessReview Nano-Aptasensing in Mycotoxin Analysis: Recent Updates and Progress
Toxins 2017, 9(11), 349; doi:10.3390/toxins9110349
Received: 22 September 2017 / Revised: 24 October 2017 / Accepted: 24 October 2017 / Published: 28 October 2017
PDF Full-text (1824 KB) | HTML Full-text | XML Full-text
Abstract
Recent years have witnessed an overwhelming integration of nanomaterials in the fabrication of biosensors. Nanomaterials have been incorporated with the objective to achieve better analytical figures of merit in terms of limit of detection, linear range, assays stability, low production cost, etc. Nanomaterials
[...] Read more.
Recent years have witnessed an overwhelming integration of nanomaterials in the fabrication of biosensors. Nanomaterials have been incorporated with the objective to achieve better analytical figures of merit in terms of limit of detection, linear range, assays stability, low production cost, etc. Nanomaterials can act as immobilization support, signal amplifier, mediator and artificial enzyme label in the construction of aptasensors. We aim in this work to review the recent progress in mycotoxin analysis. This review emphasizes on the function of the different nanomaterials in aptasensors architecture. We subsequently relate their features to the analytical performance of the given aptasensor towards mycotoxins monitoring. In the same context, a critically analysis and level of success for each nano-aptasensing design will be discussed. Finally, current challenges in nano-aptasensing design for mycotoxin analysis will be highlighted. Full article
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Figure 1

Open AccessReview Recent Advances in Mycotoxin Determination for Food Monitoring via Microchip
Toxins 2017, 9(10), 324; doi:10.3390/toxins9100324
Received: 12 September 2017 / Revised: 30 September 2017 / Accepted: 9 October 2017 / Published: 14 October 2017
PDF Full-text (1813 KB) | HTML Full-text | XML Full-text
Abstract
Mycotoxins are one of the main factors impacting food safety. Mycotoxin contamination has threatened the health of humans and animals. Conventional methods for the detection of mycotoxins are gas chromatography (GC) or liquid chromatography (LC) coupled with mass spectrometry (MS), or enzyme-linked immunosorbent
[...] Read more.
Mycotoxins are one of the main factors impacting food safety. Mycotoxin contamination has threatened the health of humans and animals. Conventional methods for the detection of mycotoxins are gas chromatography (GC) or liquid chromatography (LC) coupled with mass spectrometry (MS), or enzyme-linked immunosorbent assay (ELISA). However, all these methods are time-consuming, require large-scale instruments and skilled technicians, and consume large amounts of hazardous regents and solvents. Interestingly, a microchip requires less sample consumption and short analysis time, and can realize the integration, miniaturization, and high-throughput detection of the samples. Hence, the application of a microchip for the detection of mycotoxins can make up for the deficiency of the conventional detection methods. This review focuses on the application of a microchip to detect mycotoxins in foods. The toxicities of mycotoxins and the materials of the microchip are firstly summarized in turn. Then the application of a microchip that integrates various kinds of detection methods (optical, electrochemical, photo-electrochemical, and label-free detection) to detect mycotoxins is reviewed in detail. Finally, challenges and future research directions in the development of a microchip to detect mycotoxins are previewed. Full article
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Open AccessFeature PaperArticle Determination of Ochratoxin A in Rye and Rye-Based Products by Fluorescence Polarization Immunoassay
Toxins 2017, 9(10), 305; doi:10.3390/toxins9100305
Received: 14 September 2017 / Revised: 21 September 2017 / Accepted: 22 September 2017 / Published: 26 September 2017
PDF Full-text (809 KB) | HTML Full-text | XML Full-text
Abstract
A rapid fluorescence polarization immunoassay (FPIA) was optimized and validated for the determination of ochratoxin A (OTA) in rye and rye crispbread. Samples were extracted with a mixture of acetonitrile/water (60:40, v/v) and purified by SPE-aminopropyl column clean-up before performing
[...] Read more.
A rapid fluorescence polarization immunoassay (FPIA) was optimized and validated for the determination of ochratoxin A (OTA) in rye and rye crispbread. Samples were extracted with a mixture of acetonitrile/water (60:40, v/v) and purified by SPE-aminopropyl column clean-up before performing the FPIA. Overall mean recoveries were 86 and 95% for spiked rye and rye crispbread with relative standard deviations lower than 6%. Limits of detection (LOD) of the optimized FPIA was 0.6 μg/kg for rye and rye crispbread, respectively. Good correlations (r > 0.977) were observed between OTA contents in contaminated samples obtained by FPIA and high-performance liquid chromatography (HPLC) with immunoaffinity cleanup used as reference method. Furthermore, single laboratory validation and small-scale collaborative trials were carried out for the determination of OTA in rye according to Regulation 519/2014/EU laying down procedures for the validation of screening methods. The precision profile of the method, cut-off level and rate of false suspect results confirm the satisfactory analytical performances of assay as a screening method. These findings show that the optimized FPIA is suitable for high-throughput screening, and permits reliable quantitative determination of OTA in rye and rye crispbread at levels that fall below the EU regulatory limits. Full article
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Open AccessArticle Evaluation of Ochratoxin Recognition by Peptides Using Explicit Solvent Molecular Dynamics
Toxins 2017, 9(5), 164; doi:10.3390/toxins9050164
Received: 31 January 2017 / Revised: 9 April 2017 / Accepted: 9 May 2017 / Published: 13 May 2017
Cited by 2 | PDF Full-text (1171 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Biosensing platforms based on peptide recognition provide a cost-effective and stable alternative to antibody-based capture and discrimination of ochratoxin-A (OTA) vs. ochratoxin-B (OTB) in monitoring bioassays. Attempts to engineer peptides with improved recognition efficacy require thorough structural and thermodynamic characterization of the binding-competent
[...] Read more.
Biosensing platforms based on peptide recognition provide a cost-effective and stable alternative to antibody-based capture and discrimination of ochratoxin-A (OTA) vs. ochratoxin-B (OTB) in monitoring bioassays. Attempts to engineer peptides with improved recognition efficacy require thorough structural and thermodynamic characterization of the binding-competent conformations. Classical molecular dynamics (MD) approaches alone do not provide a thorough assessment of a peptide’s recognition efficacy. In this study, in-solution binding properties of four different peptides, a hexamer (SNLHPK), an octamer (CSIVEDGK), NFO4 (VYMNRKYYKCCK), and a 13-mer (GPAGIDGPAGIRC), which were previously generated for OTA-specific recognition, were evaluated using an advanced MD simulation approach involving accelerated configurational search and predictive modeling. Peptide configurations relevant to ochratoxin binding were initially generated using biased exchange metadynamics and the dynamic properties associated with the in-solution peptide–ochratoxin binding were derived from Markov State Models. Among the various peptides, NFO4 shows superior in-solution OTA sensing and also shows superior selectivity for OTA vs. OTB due to the lower penalty associated with solvating its bound complex. Advanced MD approaches provide structural and energetic insights critical to the hapten-specific recognition to aid the engineering of peptides with better sensing efficacies. Full article
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Open AccessArticle A Simple and Specific Noncompetitive ELISA Method for HT-2 Toxin Detection
Toxins 2017, 9(4), 145; doi:10.3390/toxins9040145
Received: 24 March 2017 / Revised: 13 April 2017 / Accepted: 14 April 2017 / Published: 20 April 2017
Cited by 2 | PDF Full-text (2069 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
We developed an HT-2 toxin-specific simple ELISA format with a positive read-out. The assay is based on an anti-immune complex (IC) scFv antibody fragment, which is genetically fused with alkaline phosphatase (AP). The anti-IC antibody specifically recognizes the IC between a primary anti-HT-2
[...] Read more.
We developed an HT-2 toxin-specific simple ELISA format with a positive read-out. The assay is based on an anti-immune complex (IC) scFv antibody fragment, which is genetically fused with alkaline phosphatase (AP). The anti-IC antibody specifically recognizes the IC between a primary anti-HT-2 toxin Fab fragment and an HT-2 toxin molecule. In the IC ELISA format, the sample is added together with the scFv-AP antibody to the ELISA plate coated with the primary antibody. After 15 min of incubation and a washing step, the ELISA response is read. A competitive ELISA including only the primary antibody recognizes both HT-2 and T-2 toxins. The anti-IC antibody makes the assay specific for HT-2 toxin, and the IC ELISA is over 10 times more sensitive compared to the competitive assay. Three different naturally contaminated matrices: wheat, barley and oats, were used to evaluate the assay performance with real samples. The corresponding limits of detection were 0.3 ng/mL (13 µg/kg), 0.1 ng/mL (4 µg/kg) and 0.3 ng/mL (16 µg/kg), respectively. The IC ELISA can be used for screening HT-2 toxin specifically and in relevant concentration ranges from all three tested grain matrices. Full article
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2016

Jump to: 2017, 2015

Open AccessArticle Comparison of In-Solution Biorecognition Properties of Aptamers against Ochratoxin A
Toxins 2016, 8(11), 336; doi:10.3390/toxins8110336
Received: 5 October 2016 / Revised: 2 November 2016 / Accepted: 8 November 2016 / Published: 15 November 2016
Cited by 3 | PDF Full-text (2371 KB) | HTML Full-text | XML Full-text
Abstract
Ochratoxin A (OTA) is a mycotoxin produced as a secondary metabolite by several species of Aspergillus and Penicillium and frequently found as a natural contaminant in a wide range of food commodities. Novel and robust biorecognition agents for detecting this molecule are required.
[...] Read more.
Ochratoxin A (OTA) is a mycotoxin produced as a secondary metabolite by several species of Aspergillus and Penicillium and frequently found as a natural contaminant in a wide range of food commodities. Novel and robust biorecognition agents for detecting this molecule are required. Aptamers are artificial nucleic acid ligands able to bind with high affinity and specificity to a given target molecule. In the last few years, three separate research groups have selected aptamers for ochratoxin A. While each of these three families of aptamers have been incorporated into various methods for detecting OTA, it is unclear if each aptamer candidate is better suited for a particular application. Here, we perform the first head-to-head comparison of solution-based binding parameters for these groups of aptamers. Based on our results, we provide recommendations for the appropriate choice of aptamer for incorporation into solution-based biorecognition assays and applications. Full article
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Open AccessReview Mycotoxin Determination in Foods Using Advanced Sensors Based on Antibodies or Aptamers
Toxins 2016, 8(8), 239; doi:10.3390/toxins8080239
Received: 25 September 2015 / Revised: 8 July 2016 / Accepted: 8 July 2016 / Published: 12 August 2016
Cited by 6 | PDF Full-text (809 KB) | HTML Full-text | XML Full-text
Abstract
Mycotoxin contamination threatens health and life of humans and animals throughout the food supply chains. Many of the mycotoxins have been proven to be carcinogens, teratogens and mutagens. The reliable and sensitive sensing methods are requested to monitor mycotoxin contamination. Advanced sensors based
[...] Read more.
Mycotoxin contamination threatens health and life of humans and animals throughout the food supply chains. Many of the mycotoxins have been proven to be carcinogens, teratogens and mutagens. The reliable and sensitive sensing methods are requested to monitor mycotoxin contamination. Advanced sensors based on antibodies or aptamers boast the advantages of high sensitivity and rapidity, and have been used in the mycotoxin sensing. These sensors are miniaturized, thereby lowering costs, and are applicable to high-throughput modes. In this work, the latest developments in sensing strategies for mycotoxin determination were critically discussed. Optical and electrochemical sensing modes were compared. The sensing methods for single mycotoxin or multiple mycotoxins in food samples were reviewed, along with the challenges and the future of antibody or aptamer-based sensors. This work might promote academic studies and industrial applications for mycotoxin sensing. Full article
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Open AccessReview Advances in Biosensors, Chemosensors and Assays for the Determination of Fusarium Mycotoxins
Toxins 2016, 8(6), 161; doi:10.3390/toxins8060161
Received: 8 April 2016 / Revised: 7 May 2016 / Accepted: 16 May 2016 / Published: 24 May 2016
Cited by 4 | PDF Full-text (283 KB) | HTML Full-text | XML Full-text
Abstract
The contaminations of Fusarium mycotoxins in grains and related products, and the exposure in human body are considerable concerns in food safety and human health worldwide. The common Fusarium mycotoxins include fumonisins, T-2 toxin, deoxynivalenol and zearalenone. For this reason, simple, fast and
[...] Read more.
The contaminations of Fusarium mycotoxins in grains and related products, and the exposure in human body are considerable concerns in food safety and human health worldwide. The common Fusarium mycotoxins include fumonisins, T-2 toxin, deoxynivalenol and zearalenone. For this reason, simple, fast and sensitive analytical techniques are particularly important for the screening and determination of Fusarium mycotoxins. In this review, we outlined the related advances in biosensors, chemosensors and assays based on the classical and novel recognition elements such as antibodies, aptamers and molecularly imprinted polymers. Application to food/feed commodities, limit and time of detection were also discussed. Full article
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Open AccessArticle Monoclonal IgA Antibodies for Aflatoxin Immunoassays
Toxins 2016, 8(5), 148; doi:10.3390/toxins8050148
Received: 23 January 2016 / Revised: 23 March 2016 / Accepted: 24 March 2016 / Published: 12 May 2016
Cited by 2 | PDF Full-text (1744 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Antibody based techniques are widely used for the detection of aflatoxins which are potent toxins with a high rate of occurrence in many crops. We developed a murine monoclonal antibody of immunoglobulin A (IgA) isotype with a strong binding affinity to aflatoxin B1
[...] Read more.
Antibody based techniques are widely used for the detection of aflatoxins which are potent toxins with a high rate of occurrence in many crops. We developed a murine monoclonal antibody of immunoglobulin A (IgA) isotype with a strong binding affinity to aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), aflatoxin G2 (AFG2) and aflatoxin M1 (AFM1). The antibody was effectively used in immunoaffinity column (IAC) and ELISA kit development. The performance of the IACs was compatible with AOAC performance standards for affinity columns (Test Method: AOAC 991.31). The total binding capacity of the IACs containing our antibody was 111 ng, 70 ng, 114 ng and 73 ng for AFB1, AFB2, and AFG1 andAFG2, respectively. Furthermore, the recovery rates of 5 ng of each AF derivative loaded to the IACs were determined as 104.9%, 82.4%, 85.5% and 70.7% for AFB1, AFB2, AFG1 and AFG2, respectively. As for the ELISA kit developed using non-oriented, purified IgA antibody, we observed a detection range of 2–50 µg/L with 40 min total test time. The monoclonal antibody developed in this research is hitherto the first presentation of quadruple antigen binding IgA monoclonal antibodies in mycotoxin analysis and also the first study of their utilization in ELISA and IACs. IgA antibodies are valuable alternatives for immunoassay development, in terms of both sensitivity and ease of preparation, since they do not require any orientation effort. Full article
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Open AccessFeature PaperArticle Comparison of Enzyme-Linked Immunosorbent Assay, Surface Plasmon Resonance and Biolayer Interferometry for Screening of Deoxynivalenol in Wheat and Wheat Dust
Toxins 2016, 8(4), 103; doi:10.3390/toxins8040103
Received: 5 February 2016 / Revised: 28 March 2016 / Accepted: 30 March 2016 / Published: 11 April 2016
Cited by 2 | PDF Full-text (922 KB) | HTML Full-text | XML Full-text
Abstract
A sample preparation method was developed for the screening of deoxynivalenol (DON) in wheat and wheat dust. Extraction was carried out with water and was successful due to the polar character of DON. For detection, an enzyme-linked immunosorbent assay (ELISA) was compared to
[...] Read more.
A sample preparation method was developed for the screening of deoxynivalenol (DON) in wheat and wheat dust. Extraction was carried out with water and was successful due to the polar character of DON. For detection, an enzyme-linked immunosorbent assay (ELISA) was compared to the sensor-based techniques of surface plasmon resonance (SPR) and biolayer interferometry (BLI) in terms of sensitivity, affinity and matrix effect. The matrix effects from wheat and wheat dust using SPR were too high to further use this screenings method. The preferred ELISA and BLI methods were validated according to the criteria established in Commission Regulation 519/2014/EC and Commission Decision 2002/657/EC. A small survey was executed on 16 wheat lots and their corresponding dust samples using the validated ELISA method. A linear correlation (r = 0.889) was found for the DON concentration in dust versus the DON concentration in wheat (LOD wheat: 233 μg/kg, LOD wheat dust: 458 μg/kg). Full article
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2015

Jump to: 2017, 2016

Open AccessArticle Development of an Enzyme-Linked Immunosorbent Assay Method Specific for the Detection of G-Group Aflatoxins
Toxins 2016, 8(1), 5; doi:10.3390/toxins8010005
Received: 31 August 2015 / Revised: 9 December 2015 / Accepted: 11 December 2015 / Published: 28 December 2015
Cited by 1 | PDF Full-text (1484 KB) | HTML Full-text | XML Full-text
Abstract
To detect and monitor G-group aflatoxins in agricultural products, we generated class-specific monoclonal antibodies that specifically recognized aflatoxins G1 and G2. Of the final three positive and stable hybridomas obtained, clone 2G6 produced a monoclonal antibody that had equal sensitivity
[...] Read more.
To detect and monitor G-group aflatoxins in agricultural products, we generated class-specific monoclonal antibodies that specifically recognized aflatoxins G1 and G2. Of the final three positive and stable hybridomas obtained, clone 2G6 produced a monoclonal antibody that had equal sensitivity to aflatoxins G1 and G2, and did not cross-react with aflatoxins B1, B2, or M1. Its IC50 values for aflatoxins G1 and G2 were 17.18 ng·mL−1 and 19.75 ng·mL−1, respectively. Using this new monoclonal antibody, we developed a competitive indirect enzyme-linked immunosorbent assay (CI-ELISA); the method had a limit of detection of 0.06 ng·mL−1. To validate this CI-ELISA, we spiked uncontaminated peanut samples with various amounts of aflatoxins G1 and G2 and compared recovery rates with those determined by a standard HPLC method. The recovery rates of the CI-ELISA ranging from 94% to 103% were comparable to those of the HPLC (92% to 102%). We also used both methods to determine the amounts of G-group aflatoxins in five peanut samples contaminated by aflatoxin B1-positive, and their relative standard deviations ranged from 8.4% to 17.7% (under 20%), which demonstrates a good correlation between the two methods. We further used this CI-ELISA to assess the ability of 126 fungal strains isolated from peanuts or field soils to produce G-group aflatoxins. Among these, seven stains producing different amounts of G-group aflatoxins were identified. Our results showed that the monoclonal antibody 2 G6-based CI-ELISA was suitable for the detection of G-group aflatoxins present in peanuts and also those produced by fungi. Full article
Open AccessArticle Highly Sensitive Colorimetric Detection of Ochratoxin A by a Label-Free Aptamer and Gold Nanoparticles
Toxins 2015, 7(12), 5377-5385; doi:10.3390/toxins7124883
Received: 16 October 2015 / Revised: 30 November 2015 / Accepted: 1 December 2015 / Published: 10 December 2015
Cited by 8 | PDF Full-text (3621 KB) | HTML Full-text | XML Full-text
Abstract
A label-free aptamer-based assay for the highly sensitive and specific detection of Ochratoxin A (OTA) was developed using a cationic polymer and gold nanoparticles (AuNPs). The OTA aptamer was used as a recognition element for the colorimetric detection of OTA based on the
[...] Read more.
A label-free aptamer-based assay for the highly sensitive and specific detection of Ochratoxin A (OTA) was developed using a cationic polymer and gold nanoparticles (AuNPs). The OTA aptamer was used as a recognition element for the colorimetric detection of OTA based on the aggregation of AuNPs by the cationic polymer. By spectroscopic quantitative analysis, the colorimetric assay could detect OTA down to 0.009 ng/mL with high selectivity in the presence of other interfering toxins. This study offers a new alternative in visual detection methods that is rapid and sensitive for OTA detection. Full article
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Open AccessArticle A Magnetic Nanoparticle Based Enzyme-Linked Immunosorbent Assay for Sensitive Quantification of Zearalenone in Cereal and Feed Samples
Toxins 2015, 7(10), 4216-4231; doi:10.3390/toxins7104216
Received: 11 September 2015 / Revised: 6 October 2015 / Accepted: 13 October 2015 / Published: 20 October 2015
Cited by 9 | PDF Full-text (698 KB) | HTML Full-text | XML Full-text
Abstract
A novel enzyme-linked immunosorbent assay based on magnetic nanoparticles and biotin/streptavidin-HRP (MNP-bsELISA) was developed for rapid and sensitive detection of zearalenone (ZEN). The detection signal was enhanced and the sensitivity of the assay was improved by combined use of antibody-conjugated magnetic nanoparticles and
[...] Read more.
A novel enzyme-linked immunosorbent assay based on magnetic nanoparticles and biotin/streptavidin-HRP (MNP-bsELISA) was developed for rapid and sensitive detection of zearalenone (ZEN). The detection signal was enhanced and the sensitivity of the assay was improved by combined use of antibody-conjugated magnetic nanoparticles and biotin-streptavidin system. Under the optimized conditions, the regression equation for quantification of ZEN was y = −0.4287x + 0.3132 (R2 = 0.9904). The working range was 0.07–2.41 ng/mL. The detection limit was 0.04 ng/mL and IC50 was 0.37 ng/mL. The recovery rates of intra-assay and inter-assay ranged from 92.8%–111.9% and 91.7%–114.5%, respectively, in spiked corn samples. Coefficients of variation were less than 10% in both cases. Parallel analysis of cereal and feed samples showed good correlation between MNP-bsELISA and liquid chromatograph-tandem mass spectrometry (R2 = 0.9283). We conclude that this method is suitable for rapid detection of zearalenone in cereal and feed samples in relevant laboratories. Full article
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Open AccessArticle Development and Evaluation of Monoclonal Antibodies for Paxilline
Toxins 2015, 7(10), 3903-3915; doi:10.3390/toxins7103903
Received: 14 August 2015 / Revised: 8 September 2015 / Accepted: 22 September 2015 / Published: 25 September 2015
Cited by 4 | PDF Full-text (994 KB) | HTML Full-text | XML Full-text
Abstract
Paxilline (PAX) is a tremorgenic mycotoxin that has been found in perennial ryegrass infected with Acremonium lolii. To facilitate screening for this toxin, four murine monoclonal antibodies (mAbs) were developed. In competitive indirect enzyme-linked immunosorbent assays (CI-ELISAs) the concentrations of PAX required
[...] Read more.
Paxilline (PAX) is a tremorgenic mycotoxin that has been found in perennial ryegrass infected with Acremonium lolii. To facilitate screening for this toxin, four murine monoclonal antibodies (mAbs) were developed. In competitive indirect enzyme-linked immunosorbent assays (CI-ELISAs) the concentrations of PAX required to inhibit signal development by 50% (IC50s) ranged from 1.2 to 2.5 ng/mL. One mAb (2-9) was applied to the detection of PAX in maize silage. The assay was sensitive to the effects of solvents, with 5% acetonitrile or 20% methanol causing a two-fold or greater increase in IC50. For analysis of silage samples, extracts were cleaned up by adsorbing potential matrix interferences onto a solid phase extraction column. The non-retained extract was then diluted with buffer to reduce solvent content prior to assay. Using this method, the limit of detection for PAX in dried silage was 15 µg/kg and the limit of quantification was 90 µg/kg. Recovery from samples spiked over the range of 100 to 1000 µg/kg averaged 106% ± 18%. The assay was applied to 86 maize silage samples, with many having detectable, but none having quantifiable, levels of PAX. The results suggest the CI-ELISA can be applied as a sensitive technique for the screening of PAX in maize silage. Full article
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Open AccessArticle Titanium Dioxide Nanoparticles (TiO2) Quenching Based Aptasensing Platform: Application to Ochratoxin A Detection
Toxins 2015, 7(9), 3771-3784; doi:10.3390/toxins7093771
Received: 30 August 2015 / Revised: 15 September 2015 / Accepted: 16 September 2015 / Published: 22 September 2015
Cited by 5 | PDF Full-text (1637 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
We demonstrate for the first time, the development of titanium dioxide nanoparticles (TiO2) quenching based aptasensing platform for detection of target molecules. TiO2 quench the fluorescence of FAM-labeled aptamer (fluorescein labeled aptamer) upon the non-covalent adsorption of fluorescent labeled aptamer
[...] Read more.
We demonstrate for the first time, the development of titanium dioxide nanoparticles (TiO2) quenching based aptasensing platform for detection of target molecules. TiO2 quench the fluorescence of FAM-labeled aptamer (fluorescein labeled aptamer) upon the non-covalent adsorption of fluorescent labeled aptamer on TiO2 surface. When OTA interacts with the aptamer, it induced aptamer G-quadruplex complex formation, weakens the interaction between FAM-labeled aptamer and TiO2, resulting in fluorescence recovery. As a proof of concept, an assay was employed for detection of Ochratoxin A (OTA). At optimized experimental condition, the obtained limit of detection (LOD) was 1.5 nM with a good linearity in the range 1.5 nM to 1.0 µM for OTA. The obtained results showed the high selectivity of assay towards OTA without interference to structurally similar analogue Ochratoxin B (OTB). The developed aptamer assay was evaluated for detection of OTA in beer sample and recoveries were recorded in the range from 94.30%–99.20%. Analytical figures of the merits of the developed aptasensing platform confirmed its applicability to real samples analysis. However, this is a generic aptasensing platform and can be extended for detection of other toxins or target analyte. Full article
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Open AccessArticle Aflatoxin B1 Detection Using a Highly-Sensitive Molecularly-Imprinted Electrochemical Sensor Based on an Electropolymerized Metal Organic Framework
Toxins 2015, 7(9), 3540-3553; doi:10.3390/toxins7093540
Received: 8 July 2015 / Revised: 10 August 2015 / Accepted: 25 August 2015 / Published: 7 September 2015
Cited by 5 | PDF Full-text (1923 KB) | HTML Full-text | XML Full-text
Abstract
A sensitive electrochemical molecularly-imprinted sensor was developed for the detection of aflatoxin B1 (AFB1), by electropolymerization of p-aminothiophenol-functionalized gold nanoparticles in the presence of AFB1 as a template molecule. The extraction of the template leads to the formation of cavities that are
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A sensitive electrochemical molecularly-imprinted sensor was developed for the detection of aflatoxin B1 (AFB1), by electropolymerization of p-aminothiophenol-functionalized gold nanoparticles in the presence of AFB1 as a template molecule. The extraction of the template leads to the formation of cavities that are able to specifically recognize and bind AFB1 through π-π interactions between AFB1 molecules and aniline moities. The performance of the developed sensor for the detection of AFB1 was investigated by linear sweep voltammetry using a hexacyanoferrate/hexacyanoferrite solution as a redox probe, the electron transfer rate increasing when the concentration of AFB1 increases, due to a p-doping effect. The molecularly-imprinted sensor exhibits a broad linear range, between 3.2 fM and 3.2 µM, and a quantification limit of 3 fM. Compared to the non-imprinted sensor, the imprinting factor was found to be 10. Selectivity studies were also performed towards the binding of other aflatoxins and ochratoxin A, proving good selectivity. Full article
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Open AccessArticle Development of a Monoclonal Antibody-Based icELISA for the Detection of Ustiloxin B in Rice False Smut Balls and Rice Grains
Toxins 2015, 7(9), 3481-3496; doi:10.3390/toxins7093481
Received: 4 July 2015 / Revised: 14 August 2015 / Accepted: 18 August 2015 / Published: 28 August 2015
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Abstract
Rice false smut is an emerging and economically-important rice disease caused by infection by the fungal pathogen Villosiclava virens. Ustiloxin B is an antimitotic cyclopeptide mycotoxin isolated from the rice false smut balls that formed in the pathogen-infected rice spikelets. A monoclonal
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Rice false smut is an emerging and economically-important rice disease caused by infection by the fungal pathogen Villosiclava virens. Ustiloxin B is an antimitotic cyclopeptide mycotoxin isolated from the rice false smut balls that formed in the pathogen-infected rice spikelets. A monoclonal antibody (mAb) designated as mAb 1B5A10 was generated with ustiloxin B—ovalbumin conjugate. A highly-sensitive and specific indirect competitive enzyme-linked immunosorbent assay (icELISA) was then developed. The median inhibitory concentration (IC50) of the icELISA was 18.0 ng/mL for the detection of ustiloxin B; the limit of detection was 0.6 ng/mL, and the calibration range was from 2.5 to 107.4 ng/mL. The LOD/LOQ values of the developed ELISA used for the determination of ustiloxin B in rice false smut balls and rice grains were 12/50 μg/g and 30/125 ng/g, respectively. The mAb 1B5A10 cross-reacted with ustiloxin A at 13.9% relative to ustiloxin B. Average recoveries of ustiloxin B ranged from 91.3% to 105.1% for rice false smut balls at spiking levels of 0.2 to 3.2 mg/g and from 92.6% to 103.5% for rice grains at spiking levels of 100 to 5000 ng/g. Comparison of ustiloxin B content in rice false smut balls and rice grains detected by both icELISA and high performance liquid chromatography (HPLC) demonstrated that the developed icELISA can be employed as an effective and accurate method for the detection of ustiloxin B in rice false smut balls, as well as rice food and feed samples. Full article
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