scholarly journals Aflatoxigenic and ochratoxigenic moulds and their toxins in tree nuts on Serbian market

2020 ◽  
pp. 9-18
Author(s):  
Irena Rakic ◽  
Gordana Dimic ◽  
Marija Skrinjar ◽  
Suncica Kocic-Tanackov
Keyword(s):  
Aspergillus Niger ◽  
Ochratoxin A ◽  
Immunosorbent Assay ◽  
Alternaria Alternata ◽  
Rhizopus Oryzae ◽  
Limit Of Detection ◽  
Average Concentration ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mean Values ◽  
Tree Nuts

In this study, moulds and mycotoxins presence in different tree nuts were investigated. The results showed that all of the 25 samples were contaminated with moulds. Mean values of total mould count varied from 1-4.9 cfu per grain. The most frequent species in hazelnut samples were Rhizopus oryzae (32.2%) and Aspergillus niger (28.9%). In walnuts A. niger (75.6%), in cashews also A. niger (42.4%) while in pistachio samples Alternaria alternata (20.7%), and Cladosporium cladosporioides (20.7%) were the most dominant. Rhizopus oligosporus was the only identified species in all almond samples (100%). Using Enzyme Linked Immunosorbent Assay (ELISA), the presence of total aflatoxins and ochratoxin A was examinated. In all analyzed samples, levels of ochratoxin A were below the limit of detection. Total aflatoxins were detected only in walnut samples with average concentration of 7.1 ?g/kg.

scholarly journals Nanobody-Alkaline Phosphatase Fusion Protein-Based Enzyme-Linked Immunosorbent Assay for One-Step Detection of Ochratoxin A in Rice

Sensors ◽  
2018 ◽  
Vol 18 (11) ◽  
pp. 4044 ◽  
Author(s):  
Zhichang Sun ◽  
Xuerou Wang ◽  
Qi Chen ◽  
Yonghuan Yun ◽  
Zongwen Tang ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Fusion Protein ◽  
Ochratoxin A ◽  
Immunosorbent Assay ◽  
Limit Of Detection ◽  
Binding Capacity ◽  
Cross Reactivity ◽  
Solvent Tolerance ◽  
Enzyme Linked Immunosorbent Assay ◽  
One Step

Ochratoxin A (OTA) has become one a focus of public concern because of its multiple toxic effects and widespread contamination. To monitor OTA in rice, a sensitive, selective, and one-step enzyme-linked immunosorbent assay (ELISA) using a nanobody-alkaline phosphatase fusion protein (Nb28-AP) was developed. The Nb28-AP was produced by auto-induction expression and retained an intact antigen-binding capacity and enzymatic activity. It exhibited high thermal stability and organic solvent tolerance. Under the optimal conditions, the developed assay for OTA could be finished in 20 min with a half maximal inhibitory concentration of 0.57 ng mL−1 and a limit of detection of 0.059 ng mL−1, which was 1.1 times and 2.7 times lower than that of the unfused Nb28-based ELISA. The Nb28-AP exhibited a low cross-reactivity (CR) with ochratoxin B (0.92%) and ochratoxin C (6.2%), and an ignorable CR (<0.10%) with other mycotoxins. The developed Nb-AP-based one-step ELISA was validated and compared with a liquid chromatography-tandem mass spectrometry method. The results show the reliability of Nb-AP-based one-step ELISA for the detection of OTA in rice.

scholarly journals Co-occurrence of Aflatoxins, Ochratoxin A, Fumonisins, and Zearalenone in Cereals and Feed, Determined by Competitive Direct Enzyme-Linked Immunosorbent Assay and Thin-Layer Chromatography

2009 ◽  
Vol 60 (4) ◽  
pp. 427-434 ◽  
Author(s):  
Maja Klarić ◽  
Zdenka Cvetnić ◽  
Stjepan Pepeljnjak ◽  
Ivan Kosalec
Keyword(s):  
Thin Layer ◽  
Thin Layer Chromatography ◽  
Ochratoxin A ◽  
Immunosorbent Assay ◽  
Limit Of Detection ◽  
Permissible Limit ◽  
Enzyme Linked Immunosorbent Assay ◽  
Wide Range ◽  
Food And Feed ◽  
Layer Chromatography

Co-occurrence of Aflatoxins, Ochratoxin A, Fumonisins, and Zearalenone in Cereals and Feed, Determined by Competitive Direct Enzyme-Linked Immunosorbent Assay and Thin-Layer ChromatographyAspergillus, Penicillium, andFusariumspecies frequently contaminate crops. For this reason mycotoxins such as aflatoxins (AFs), ochratoxin A (OTA), fumonisins (FBs), and zearalenone (ZEA) are found in food and feed in a wide range of concentrations, depending on environmental and storage conditions. Consumption of mycotoxin-contaminated food and feed has been associated with acute and chronic poisoning and carcinoma. The aim of this study was to determine the incidence and co-occurrence of AFs (B1+B2+G1+G2), OTA, FBs (B1+B2+B3), and ZEA in 37 samples of cereals and feed randomly collected in 2007 from households of an endemic nephropathy (EN) area in Croatia. The mycotoxins were determined using the competitive direct ELISA test (CD-ELISA) in combination with thin-layer chromatography (TLC). The most frequent mycotoxin was ZEA (92%, mean 318.3 μg kg-1), followed by FBs (27%, 3690 μg kg-1), AFs (24.3%, 4.6 μg kg-1), and OTA (16.2%, 9.8 μg kg-1). Levels of AFs, ZEA, and FBs detected by CD-ELISA significantly correlated with the TLC results. However, only one OTA-positive sample was confirmed by TLC due to its high limit of detection. The levels of these mycotoxins were below the permissible limit for animal feed. Twenty-nine percent of cereals were contaminated with FBs, OTA, or ZEA in mass fractions above the permissible limit for humans. Co-occurrence of two toxins varied between 4.2% and 54% and of three between 4.2% and 7.6%. Prolonged co-exposure to AFs, OTA, FBs, and ZEA might increase the risk of various chronic diseases.

SENSITIVE AND SPECIFIC ENZYME-LINKED IMMUNOSORBENT ASSAY FOR UROKINASE-TYPE PLASMINOGEN ACTIVATOR IN HUMAN PLASMA

1987 ◽  
Author(s):  
J Grøndahl-HANSEN ◽  
N Agerlin ◽  
L S Nielsen ◽  
K Danø
Keyword(s):  
Plasminogen Activator ◽  
Immunosorbent Assay ◽  
Enzyme Linked Immunosorbent Assay ◽  
Mean Values ◽  
Amino Terminal ◽  
Type Plasminogen Activator ◽  
Healthy Donors ◽  
Urokinase Type Plasminogen Activator ◽  
The Mean

An enzyme-linked immunosorbent assay (ELISA) was developed for the measurement of human urokinase-type plasminogen activator (u-PA) in plasma and serum. Microtiter plates were coated with a monoclonal antibody and incubated with standard or sample. Bound u-PA was quantitated with polyclonal antibodies conjugated with biotin, followed by avidin-peroxidase. The assay was 10-fold as sensitive as other previously reported ELISAs, the detection limit being approximately 1 pg of u-PA in a volume of 100 μl with a linear dose-response up to 15 pg of u-PA. The assay detected active u-PA and its inactive proenzyme form equally well and the recovery of both forms was higher than 90% in plasma. A variety of structurally related proteins, including t-PA, were tested, but no reaction with proteins other than u-PA and its amino-terminal degradation product were observed. The intra-assay and inter-assay coefficients of variation for determination of u-PA in plasma were 7.6% and 8.4%, respectively. The assay was equally applicable to serum. The values obtained with plasma and serum were similar, and the results were not affected by small variations in the preparation of the samples. The ELISA was used to measure the concentration of u-PA in plasma from 34 healthy donors. The mean values for u-PA in plasma from healthy donors was 1.1 ng/ml ± 0.3 ng/ml (SD) (range 0.6 - 1.5 ng/ml). No significant differences were found between men and women and no correlation between u-PA concentration and age could be demonstrated.The mean u-PA concentration in plasma from healthy donors obtained in this study is substantially lower than that reported by others. This might be due to different methods of determination of the protein content of the standard preparations or to differences in the specificity of the assays.

MYCOTOXINS IN BEANS OF PEANUTS GROWN IN TAJIKISTAN

2018 ◽  
Vol 1 (4) ◽  
pp. 74-77
Author(s):  
Sh. I. Razokov ◽  
◽  
D. M. Mirzoev ◽  
G. P. Kononenko ◽  
A. A. Burkin ◽  
...  
Keyword(s):  
Ochratoxin A ◽  
Immunosorbent Assay ◽  
Cyclopiazonic Acid ◽  
Practical Significance ◽  
Enzyme Linked Immunosorbent Assay ◽  
Test Systems ◽  
The Republic ◽  
Combined Contamination

The article presents the results of an extensive mycotoxicological examination of 11 samples of peanut beans grown in two regions of the Republic of Tajikistan. The determination of 16 mycotoxins was carried out by indirect competitive enzyme-linked immunosorbent assay using commercial and certified research test systems. It has been established that for peanut beans in this area, a combined contamination by a group of sanitary-significant mycotoxins, including diacetoxyscirpenol, alternariol, ochratoxin A, PR-toxin and cyclopiazonic acid, is characteristic. The prospects of further research and the practical significance of the results are discussed.

Development of a quantitative and sensitive enzyme-linked immunosorbent assay for ochratoxin A using antibodies from the yolk of the laying hen

1993 ◽  
Vol 41 (10) ◽  
pp. 1784-1789 ◽  
Author(s):  
J. R. Clarke ◽  
R. R. Marquardt ◽  
A. Oosterveld ◽  
A. A. Frohlich ◽  
F. J. Madrid ◽  
...  
Keyword(s):  
Ochratoxin A ◽  
Immunosorbent Assay ◽  
Laying Hen ◽  
Enzyme Linked Immunosorbent Assay

scholarly journals Dramatically Enhancing the Sensitivity of Immunoassay for Ochratoxin A Detection by Cascade-Amplifying Enzyme Loading

Toxins ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 781
Author(s):  
Zhuolin Song ◽  
Lin Feng ◽  
Yuankui Leng ◽  
Mingzhu Huang ◽  
Hao Fang ◽  
...  
Keyword(s):  
Ochratoxin A ◽  
Limit Of Detection ◽  
High Sensitivity ◽  
Cross Reaction ◽  
Molar Ratio ◽  
Quantitative Detection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Enzyme Loading ◽  
M13 Bacteriophage ◽  
Mimic Peptide

Enzyme-linked immunosorbent assay (ELISA) is widely used in the routine screening of mycotoxin contamination in various agricultural and food products. Herein, a cascade-amplifying system was introduced to dramatically promote the sensitivity of an immunoassay for ochratoxin A (OTA) detection. Specifically, a biotinylated M13 bacteriophage was introduced as a biofunctional competing antigen, in which a seven-peptide OTA mimotope fused on the p3 protein of M13 was used to specifically recognize an anti-OTA monoclonal antibody, and the biotin molecules modified on capsid p8 proteins were used in loading numerous streptavidin-labeled polymeric horseradish peroxidases (HRPs). Owing to the abundance of biotinylated p8 proteins in M13 and the high molar ratio between HRP and streptavidin in streptavidin-polyHRP, the loading amount of HRP enzymes on the M13 bacteriophage were greatly boosted. Hence, the proposed method exhibited high sensitivity, with a limit of detection of 2.0 pg/mL for OTA detection, which was 250-fold lower than that of conventional ELISA. In addition, the proposed method showed a slight cross-reaction of 2.3% to OTB, a negligible cross-reaction for other common mycotoxins, and an acceptable accuracy for OTA quantitative detection in real corn samples. The practicability of the method was further confirmed with a traditional HRP-based ELISA method. In conclusion, the biotinylated bacteriophage and polyHRP structure showed potential as a cascade-amplifying enzyme loading system for ultra-trace OTA detemination, and its application can be extended to the detection of other analytes by altering specific mimic peptide sequences.

scholarly journals Identification and dietary exposure assessment of tetracycline and penicillin residues in fluid milk, yogurt, and labneh: A cross-sectional study in Lebanon

2019 ◽  
Vol 12 (4) ◽  
pp. 527-534 ◽  
Author(s):  
Suzanne Kabrite ◽  
Christelle Bou-Mitri ◽  
Jessy El Hayek Fares ◽  
Hussein F. Hassan ◽  
Jocelyne Matar Boumosleh
Keyword(s):  
Human Health ◽  
Dairy Products ◽  
Detection Rate ◽  
Limit Of Detection ◽  
Human Health Risk ◽  
Daily Intake ◽  
Mean Value ◽  
Enzyme Linked Immunosorbent Assay ◽  
Antibiotic Residues ◽  
Mean Values

Background and Aim: The safety and quality of dairy products are considered to be of significant importance to human health. Although antimicrobial drugs are essential for disease treatment in modern medicine, the use of these drugs can have undesired consequences for human and animal health. This study aimed to investigate the presence of tetracycline and penicillin residues in raw, pasteurized, and UHT cow's milk of different fat contents, as well as in the dairy products yogurt and labneh, a traditional Lebanese product. Materials and Methods: A total of 44 samples, 4 raw, 9 UHT, 9 pasteurized milk, 10 yogurt, and 12 labneh samples from common local brands available in the Lebanese market were collected from Keserwan regions in May 2016. Tetracycline and penicillin residues were determined using a competitive enzyme-linked immunosorbent assay (ELISA) technique. Results: The mean values for tetracycline and penicillin were all below the limit of detection (LOD) of the ELISA kit of a maximum standard concentration of 1.80 μg/kg and 4.00 μg/kg, respectively. All samples tested positive for antibiotic residues. The detection rate for tetracycline in milk (n=22) samples was 86.4% with a mean residues value of 1.16±0.70 μg/kg. The detection rate of tetracycline in labneh (n=12) and yogurt (n=10) samples was 50% for each with a mean value of 1.76±0.40 μg/kg and 0.63±0.12 μg/kg, respectively. As for penicillin residues, 90.9% of the milk (n=22) samples tested positive with a mean value of 0.52±0.25 μg/kg. The detection rate in labneh (n=12) and yogurt (n=10) samples was 0% for penicillin residues, where mean values were all below the LOD (<1.25 μg/kg) for these dairy products. None of the samples exceeded the maximum residue levels. The estimated dietary intake (EDI) for tetracycline and penicillin residues for all dairy products is 2.09 ng/kg body weight (BW)/day resulting in 0.007% of the acceptable daily intake (ADI) and 1.83 ng/kg BW/day resulting in 0.006% of the ADI, respectively. Conclusion: All EDI values were below the ADI set for each antibiotic residue and do not exceed relevant toxicological reference values. However, concerns might still be present from consumption of other animal food products containing residues. Moreover, the long-term exposure to such residues is still unknown as a result of bioaccumulation; it is a challenging process to determine the actual dietary consumption of foods containing antibiotic residues; hence, the human health risk cannot be easily predicted.

scholarly journals Mycotoxins as a risk in the grain food

2009 ◽  
pp. 79-86 ◽  
Author(s):  
Jovana Matic ◽  
Jasna Mastilovic ◽  
Ivana Cabarkapa ◽  
Anamarija Mandic
Keyword(s):  
European Union ◽  
Secondary Metabolites ◽  
Ochratoxin A ◽  
Immunosorbent Assay ◽  
Toxic Effects ◽  
Enzyme Linked Immunosorbent Assay ◽  
The European Union ◽  
The Mean ◽  
Grain Foods

Mycotoxins are toxic secondary metabolites of fungi that contaminate a large variety of foods and have toxic effects on humans. The best protection against mycotoxins is to monitor their presence in food. This paper shows the screening results of mycotoxins present in 76 samples of different groups of grain foods. Samples of grain food were analyzed for contamination with aflatoxins, ochratoxin A, zearalenone, fumonisins and deoxynivalenol. Analysis were conducted using competitive enzyme-linked immunosorbent assay (ELISA). None of the samples was contaminated with aflatoxins. The most predominant mycotoxin was ochratoxin A with the mean level of 4.84 ? 4.49 ppb in 19.7% of the examined samples. Zearalenone, fumonisins, and deoxynivalenol were found in 9.21, 14.5 and 3.9% of the samples, respectively. Mycotoxin content in the investigated samples was compared with the regulations of Serbia and those of the European Union.

Simulation model for predicting limit of detection and range of quantitation of competitive enzyme-linked immunosorbent assay

2007 ◽  
Vol 103 (5) ◽  
pp. 427-431 ◽  
Author(s):  
Dong Hwan Choi ◽  
Yoshio Katakura ◽  
Rieko Matsuda ◽  
Yuzuru Hayashi ◽  
Kazuaki Ninomiya ◽  
...  
Keyword(s):  
Simulation Model ◽  
Immunosorbent Assay ◽  
Limit Of Detection ◽  
Enzyme Linked Immunosorbent Assay

scholarly journals Change of Amino Acid Residues in Idiotypic Nanobodies Enhanced the Sensitivity of Competitive Enzyme Immunoassay for Mycotoxin Ochratoxin A in Cereals

Toxins ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 273
Author(s):  
Caixia Zhang ◽  
Weiqi Zhang ◽  
Xiaoqian Tang ◽  
Qi Zhang ◽  
Wen Zhang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Ochratoxin A ◽  
Limit Of Detection ◽  
Basic Knowledge ◽  
Affinity Constant ◽  
Enzyme Linked Immunosorbent Assay ◽  
Amino Acid Residues ◽  
Assay Sensitivity ◽  
Coating Antigen ◽  
Relative Standard

Anti-idiotypic nanobodies, usually expressed by gene engineering protocol, has been shown as a nontoxic coating antigen for toxic compound immunoassays. We here focused on how to increase immunoassay sensitivity by changing the nanobody’s primary sequence. In the experiments, two anti-idiotype nanobodies against monoclonal antibody 1H2, which is specific to ochratoxin A, were obtained and named as nontoxic coating antigen 1 (NCA1) and nontoxic coating antigen 2 (NCA2). Three differences between the nanobodies were discovered. First, there are six amino acid residues (AAR) of changes in the complementarity determining region (CDR), which compose the antigen-binding site. One of them locates in CDR1 (I–L), two of them in CDR2 (G–D, E–K), and three of them in CDR3 (Y–H, Y–W). Second, the affinity constant of NCA1 was tested as 1.20 × 108 L mol−1, which is about 4 times lower than that of NCA2 (5.36 × 108 L mol−1). Third, the sensitivity (50% inhibition concentration) of NCA1 for OTA was shown as 0.052 ng mL−1, which was 3.5 times lower than that of nontoxic coating antigen 2 (0.015 ng mL−1). The results indicate that the AAR changes in CDR of the anti-idiotypic nanobodies, from nonpolar to polar, increasing the affinity constant may enhance the immunoassay sensitivity. In addition, by using the nontoxic coating antigen 2 to substitute the routine synthetic toxic antigen, we established an eco-friendly and green enzyme-linked immunosorbent assay (ELISA) method for rapid detection of ochratoxin A in cereals. The half-maximal inhibitory concentration (IC50) of optimized ELISA was 0.017 ng mL−1 with a limit of detection (LOD) of 0.003 ng mL−1. The optimized immunoassay showed that the average recoveries of spiked corn, rice, and wheat were between 80% and 114.8%, with the relative standard deviation (RSD) ranging from 3.1–12.3%. Therefore, we provided not only basic knowledge on how to improve the structure of anti-idiotype nanobody for increasing assay sensitivity, but also an available eco-friendly ELISA for ochratoxin A in cereals.

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