Enzyme-Linked Immunosorbent Assay of Total Aflatoxins B1, B2, and G1 in Corn: Follow-up Collaborative Study

1994 ◽  
Vol 77 (3) ◽  
pp. 655-658 ◽  
Author(s):  
Mary W Trucksess ◽  
Michael E Stack
Keyword(s):  
Immunosorbent Assay ◽  
Polyclonal Antibodies ◽  
Collaborative Study ◽  
Screening Method ◽  
Test Sample ◽  
Enzyme Linked Immunosorbent Assay ◽  
Test Device ◽  
Total Aflatoxin ◽  
Peroxidase Conjugate

Abstract A direct competitive enzyme-linked immunosorbent assay screening method for af latoxins at 20 ng/g in corn was studied by 15 collaborating laboratories. Test samples of corn were extracted by blending with methanol-water (8 + 2). The extracts were filtered and the filtrates were diluted with buffer to a final methanol concentration of <30%. Each diluted filtrate was applied to a test device containing a filter with immobilized polyclonal antibodies specific to aflatoxins B1, B2, and G1. Aflatoxin Bi-peroxidase conjugate was added, the test device was washed with water, and a mixture of hydrogen peroxide and tetramethylbenzidine was added. A test sample was judged to contain ≥20 ng af latoxins/g when, after exactly 1 min, no color was observed on the filter; if a blue or gray color developed, the test sample was judged to contain <20 ng aflatoxins/g. All laboratories correctly identified naturally contaminated corn test samples. Only one false positive was found for controls containing no aflatoxins. The correct responses for positive test samples spiked at levels of 10,20, and 30 ng aflatoxins/g (the ratio of B1:B2:G1 was 15:1:3) were 67,97, and 100%, respectively. This method was adopted first action by AOAC INTERNATIONAL as a change in method for 990.34 for screening for aflatoxins B1, B2, and G1 in corn at total aflatoxin concentrations of ≥20 ng/g.

Enzyme-Linked Immunosorbent Assay of Aflatoxins Bi, B2, and Gi in Corn, Cottonseed, Peanuts, Peanut Butter, and Poultry Feed: Collaborative Study

1989 ◽  
Vol 72 (6) ◽  
pp. 957-962 ◽  
Author(s):  
Mary W Trucksess ◽  
Michael E Stack ◽  
Stanley Nesheim ◽  
Douglas L Park ◽  
Albert E Pohland
Keyword(s):  
Immunosorbent Assay ◽  
Polyclonal Antibodies ◽  
Peanut Butter ◽  
Collaborative Study ◽  
Screening Method ◽  
Steam Bath ◽  
Test Sample ◽  
Positive Test ◽  
Poultry Feed ◽  
Enzyme Linked Immunosorbent Assay

Abstract A direct competitive enzyme-linked immunosorbent assay (ELISA) screening method for aflatoxins at 20 ng/g was studied by 12 collaborators. Test samples of peanut butter were extracted by blending with methanol-water-hexane (55 + 45 + 100) and heating the test extracts on a steam bath; test samples of the other commodities were extracted by blending with methanol-water (80 + 20). All test extracts were filtered and the filtrates were diluted with buffer to a final methanol concentration of <30%. Each diluted filtrate was applied to a cup containing a filter with immobilized polyclonal antibodies specific to aflatoxins B1, B2, and G1. Aflatoxin B1-peroxidase conjugate was added, the cup was washed with water, and a mixture of hydrogen peroxide and tetramethylbenzidine was added. The test sample was judged to contain >20 ng aflatoxins/g when, after exactly 1 min, no color was observed on the filter; when a blue or gray color developed, the test sample was judged to contain <20 ng aflatoxins/g. All collaborators correctly identified naturally contaminated corn and raw peanut positive test samples. No false positives were found for controls containing <2 ng aflatoxins/g. The correct responses for positive test samples spiked at levels of 10, 20, and >30 ng aflatoxins/g (the ratio of B1, B2, and G1 was 10:1:3) were 52, 86, and 96%, respectively. The method, which is rapid and simple, has been adopted official first action for screening for aflatoxins at >20 ng/g in cottonseed and peanut butter and for aflatoxins at >30 ng/g in corn and raw peanuts. The procedure will require further study for poultry feed. Positive test samples may require reanalysis by an official, quantitative method.

Determination of Total Aflatoxin Levels in Peanut Butter by Enzyme-Linked Immunosorbent Assay: Collaborative Study

1992 ◽  
Vol 75 (4) ◽  
pp. 693-697 ◽  
Author(s):  
Alan L Patey ◽  
Matthew Sharman ◽  
John Gilbert
Keyword(s):  
Immunosorbent Assay ◽  
Aflatoxin B1 ◽  
Peanut Butter ◽  
Collaborative Study ◽  
The United States ◽  
Enzyme Linked Immunosorbent Assay ◽  
Total Aflatoxin ◽  
Aflatoxin Level ◽  
Relative Standard

Abstract Laboratories in Australia, Japan, Spain, the United Kingdom, and the United States participated in a collaborative study to evaluate a commercial enzyme- linked immunosorbent assay for the determination of total aflatoxin. Collaborators were sent 10 randomly numbered samples (5 blind duplicates) of roasted peanut butter. Two pairs were "blank" peanut butters to which aflatoxin B1, B2, G1, and G2 standards had been added. The other 3 pairs of peanut butters were 1 low aflatoxin level sample and 2 naturally contaminated samples. The assay is based on indirect competition. Test samples containing (free) aflatoxin, added to aflatoxin-coated microwells, compete for specific monoclonal rat anti-aflatoxin. As the concentration of aflatoxin in the test samples increases, the amount of rat antiaflatoxin binding to the aflatoxin attached to the well decreases. After a wash step to remove unbound material, the amount of rat anti-aflatoxin bound to the well is determined by its reaction with peroxidase conjugated rabbit anti-rat globulin. Bound peroxidase activity is then determined by the addition of a substrate, whose color development is inversely proportional to the aflatoxin concentration and is measured by absorbance. Coefficients of variation (CV) for total aflatoxin concentrations, for mean levels of 9,30, and 89µg/kg, were between 28 and 37% for the low level and 2 naturally contaminated samples, which contained mainly aflatoxin B1. CVs for the spiked samples were lower (24-25%) for mean levels of 11 and 20 µg/kg; recoveries were 84 and 89%, respectively. Ranges for relative standard deviations for repeatabilty and reproducibility were 9-30% and 25-37%, respectively. The method has been adopted first action by AOAC International.

scholarly journals Determination of Vegetal Proteins in Milk Powder by Enzyme-Linked Immunosorbent Assay: Interlaboratory Study

2002 ◽  
Vol 85 (6) ◽  
pp. 1390-1397 ◽  
Author(s):  
Lourdes Sánchez ◽  
María D Pérez ◽  
Pilar Puyol ◽  
Miguel Calvo ◽  
Gary Brett ◽  
...  
Keyword(s):  
Immunosorbent Assay ◽  
Polyclonal Antibodies ◽  
Collaborative Study ◽  
Milk Powder ◽  
Enzyme Linked Immunosorbent Assay ◽  
Wheat Proteins ◽  
Blind Test ◽  
Relative Standard ◽  
Standard Deviations ◽  
Phosphate Buffered Saline

Abstract Eight laboratories participated in a collaborative study to evaluate an enzyme-linked immunosorbent assay (ELISA) to determine soy, pea, and wheat proteins in pasteurized or ultra-high temperature (UHT) milk powders. To perform this assay, polyclonal antibodies for soy, pea, and wheat proteins were obtained from rabbit sera. Collaborators received calibration standards composed of milk powder containing 0–8% (w/w) vegetal protein in total protein and blind test samples containing approximately 1, 2, and 5% (w/w) vegetal protein. An indirect competitive ELISA was performed with a kit prepared by a participating laboratory; the kit contained plates coated with soy, pea, or wheat proteins, the corresponding specific antisera, enzyme-labeled second antibody, and substrate solution. Test samples and calibrants were extracted with phosphate-buffered saline, pH 7.4, containing 0.05% Tween and assayed with the ELISA kits. The degree of adulteration was affected by the type of heat treatment applied to the samples. The estimated percentage of vegetal protein addition was close to the theoretical value for pasteurized samples but much lower for UHT samples. For pasteurized samples, intralaboratory relative standard deviations ranged from 5 to 22% and interlaboratory relative standard deviations ranged from 14 to 34%.

Enzyme-Linked Immunosorbent Assay for Detection of Zearalenone in Corn, Wheat, and Pig Feed: Collaborative Study

1994 ◽  
Vol 77 (6) ◽  
pp. 1500-1508 ◽  
Author(s):  
Glenn A Bennett ◽  
Terry C Nelsen ◽  
Brjnton M Miller
Keyword(s):  
Immunosorbent Assay ◽  
Collaborative Study ◽  
Screening Method ◽  
The United States ◽  
False Positives ◽  
Enzyme Linked Immunosorbent Assay ◽  
Elisa Method ◽  
False Negatives ◽  
Relative Standard ◽  
Pig Feed

Abstract A direct competitive enzyme-linked immunosorbent assay (ELISA) screening method for zearalenone in corn, wheat, and feed at 500 ng/g was evaluated by 23 collaborators (22 laboratories) in an international collaborative study. Eighteen samples of spiked or naturally contaminated corn, wheat, and pig feed were prepared by the sponsoring laboratory and sent for testing with complete test kits to participating collaborators in Canada, Italy, Sweden, The Netherlands, and the United States. Test samples were extracted with methanolwater solution (70 + 30) by shaking on a wrist-action shaker for 3 min. A portion of the extract was mixed with an equal volume of zearalenone-enzyme conjugate, and the mixture was incubated with zearalenone-specific monoclonal antibodies coated onto microtiter wells. All test samples were assayed in duplicate. One of 52 (2%) blanks was reported positive. Thirty-nine of the 52 (75%) samples that were spiked at 500 ng/g were reported as positive. Forty-nine of the 51 (96%) samples with concentrations at or above 1000 ng/g were reported as positive. The overall incidence of false negatives was 6.0% and the incidence of false positives was 22.7% by the ELISA method. Only one (3.4%) false negative was reported for samples containing ≥800 ng/g. In the spectrophotometric method, 8 collaborators determined approximate levels of zearalenone in test samples from standard curves constructed from spiked extracts (0–3000 ng/g of each commodity tested). This method gave and overall incidence of false negatives of 5.7% and false positives of 17.8%. Average relative standard deviations, RSDr (repeatability) and RSDR (reproducibility), were 11.6 and 25.1% for spiked samples and 11.7 and 33.1% for naturally contaminated samples, respectively. Standard curves were constructed with each set of samples assayed. Comparison of absorbance values from these standard curves indicate the performance of reagents and antibody used in the assay. The ELISA method has been adopted first action by AOAC INTERNATIONAL as a screening method for zearalenone at ≥800 ng/g in corn, wheat, and pig feed.

scholarly journals LOCATE Enzyme-Linked Immunosorbent Assay for Detection of Salmonella in Food: Collaborative Study

1998 ◽  
Vol 81 (2) ◽  
pp. 419-437 ◽  
Author(s):  
Vidhya Gangar ◽  
Michael S Curiale ◽  
Armando D'onorio ◽  
Carol Donnelly ◽  
Paul Dunnigan ◽  
...  
Keyword(s):  
Collaborative Study ◽  
Screening Method ◽  
Culture Method ◽  
Microtiter Plate ◽  
Black Pepper ◽  
Soy Flour ◽  
Food Type ◽  
Enzyme Linked Immunosorbent Assay ◽  
Data Sets ◽  
Whole Egg

abstract A collaborative study was performed in 27 laboratories to validate the enzyme-linked immunosorbent procedure LOCATE for rapid detection of Salmonella in foods. Results were read visually and with a microtiter plate reader. The LOCATE method was compared with the Bacteriological Analytical Manual (BAM)/AOAC INTERNATIONAL culture method for detecting Salmonella in 6 foods: milk chocolate, nonfat dry milk, dried whole egg, soy flour, ground black pepper, and ground raw turkey. Two foods—dried whole egg and black pepper—required repeat rounds because insufficient data sets were produced initially (AOAC INTERNATIONAL stipulates a minimum of 15 sets per food type). Each laboratory tested one or more of the 6 foods. A total of 1 439 samples were analyzed, and no significant differences (P <0.05) were observed between LOCATE with either visual or reader detection and BAM/AOAC INTERNATIONAL results. The LOCATE screening method with visual or reader detection is recommended for Official First Action Approval

scholarly journals Enzyme-Linked Immunosorbent Assay for Simultaneous Detection of Two Fungicides Kresoxim-methyl and Trifloxystrobin in Oranges

2016 ◽  
Vol 34 (No. 5) ◽  
pp. 429-438 ◽  
Author(s):  
Yang Wei ◽  
Zhu Jie ◽  
Liu Ming-Qi ◽  
Dai Xian-Jun
Keyword(s):  
Immunosorbent Assay ◽  
Reaction Rate ◽  
Polyclonal Antibodies ◽  
Simultaneous Detection ◽  
Acid Methyl Ester ◽  
Enzyme Linked Immunosorbent Assay ◽  
Strobilurin Fungicides ◽  
Acid Methyl ◽  
Recovery Study ◽  
The Common

To assemble an indirect competitive enzyme-linked immunosorbent assay (ELISA) for estimating two strobilurin fungicides at the same time, the hapten was synthesised which contained the common active group, (E)-2-(2-bromo-phenyl)-2-(methoxyimino) acetic acid methyl ester (OEBr) in kresoxim-methyl and trifloxystrobin. The immunogen and coating antigen were respectively prepared through conjugating the above-mentioned hapten with BSA and OVA by the mixed anhydride and activated ester methods, and polyclonal antibodies were produced by immunised rabbits. An enzyme-linked immunosorbent assay was developed for simultaneous detection of kresoxim-methyl and trifloxystrobin. In ELISA, the antiserum showed high affinity and sensitivity to kresoxim-methyl and trifloxystrobin, and their IC<sub>50</sub> value and detection limit (expressed as IC<sub>10</sub>) were 14.7 and 0.0044 µg/ml, respectively, for kresoxim-methyl, and 22.9 and 0.014 µg/ml, respectively for trifloxystrobin. The cross-reaction rate was below 0.1% for other strobilurin fungicides. Recovery study of ELISA from spiked samples of homogenised peeled oranges (final concentrations of 100, 10, and 1 µg/ml) resulted in recovery levels in the range of 82–104%.

Quantification of serum amyloid P by enzyme-linked immunosorbent assay

1991 ◽  
Vol 37 (10) ◽  
pp. 1742-1745 ◽  
Author(s):  
R Saïle ◽  
A Kandoussi ◽  
M Deveaux ◽  
J Descamps ◽  
E Hachulla ◽  
...  
Keyword(s):  
Human Plasma ◽  
Immunosorbent Assay ◽  
Polyclonal Antibodies ◽  
Serum Amyloid ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Amyloid P ◽  
Assay Procedure ◽  
Normal Individuals ◽  
Sandwich Type ◽  
Amyloid P

Abstract This enzyme-linked immunosorbent assay procedure for quantifying serum amyloid P (SAP) in human plasma makes use of affinity-purified polyclonal antibodies to SAP in a "sandwich"-type format. The procedure is sensitive, reproducible, simple, and easily automatable. Results correlate well with those by a rocket immunoelectrophoresis method performed with the same antibodies. Sera from apparently normal individuals had a mean SAP content of 44.17 mg/L and increased with age.

Development of a Highly Sensitive Enzyme-Linked Immunosorbent Assay Based on Polyclonal Antibodies for the Detection of Polychlorinated Dibenzo-p-dioxins

1998 ◽  
Vol 70 (6) ◽  
pp. 1092-1099 ◽  
Author(s):  
Yukio Sugawara ◽  
Shirley J. Gee ◽  
James R. Sanborn ◽  
S. Douglass Gilman ◽  
Bruce D. Hammock
Keyword(s):  
Immunosorbent Assay ◽  
Polyclonal Antibodies ◽  
Enzyme Linked Immunosorbent Assay ◽  
Highly Sensitive

scholarly journals Predicting Aflatoxin and Fumonisin in Shelled Corn Lots Using Poor-Quality Grade Components

2006 ◽  
Vol 89 (2) ◽  
pp. 433-440 ◽  
Author(s):  
Anders S Johansson ◽  
Thomas B Whitaker ◽  
Winston M Hagler ◽  
Daryl T Bowman ◽  
Andy B Slate ◽  
...  
Keyword(s):  
North Carolina ◽  
Total Concentration ◽  
Screening Method ◽  
Test Sample ◽  
Poor Quality ◽  
Foreign Material ◽  
Total Aflatoxin ◽  
Quality Grade ◽  
Aflatoxin Concentration ◽  
Inspection Service

Abstract A study was conducted to determine if aflatoxin and fumonisin are concentrated in the poor-quality grade components of shelled corn. Four 1.0 kg test samples were each taken from 23 lots of shelled corn marketed in North Carolina. Inspectors from the Federal Grain Inspection Service divided each test sample into 3 grade components: (1) damaged kernels (DM), (2) broken corn and foreign material (BCFM), and )3) whole kernels (WH). The aflatoxin and fumonisin concentration was measured in each component and a mass balance equation was used to calculate the total concentration of each mycotoxin in each test sample. Averaged across all test samples, the aflatoxin concentrations in the DM, BCFM, and WH components were 1300.3, 455.2, and 37.3 ppb, respectively. Averaged across all test samples, the fumonisin concentrations in the DM, BCFM, and WH components were 148.3, 51.3, and 1.8 ppm, respectively. The DM and BCFM components combined accounted for only 5.0% of the test sample mass, but accounted for 59.8 and 77.5% of the total aflatoxin and fumonisin mass in the test sample, respectively. Both aflatoxin mass (ng) and aflatoxin concentration (ng/g) in the combined DM and BCFM components had high correlations with aflatoxin concentration in the lot. The highest correlation occurred when aflatoxin mass (ng) in the combined DM and BCFM components was related to aflatoxin concentration in the lot (0.964). Similar results were obtained for fumonisin. This study indicated that measuring either aflatoxin or fumonisin in the combined DM and BCFM grade components could be used as a screening method to predict either aflatoxin or fumonisin in a bulk lot of shelled corn.

scholarly journals Aflatoxin Plate Kit

2011 ◽  
Vol 94 (5) ◽  
pp. 1519-1530
Author(s):  
Arthur Trombley ◽  
Titan Fan ◽  
Robert LaBudde
Keyword(s):  
Immunosorbent Assay ◽  
Hplc Method ◽  
Aflatoxin Contamination ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rapid Testing ◽  
Total Aflatoxin ◽  
Standard Curve ◽  
Independent Laboratory ◽  
Test Kit

Abstract The level of total aflatoxin contamination was analyzed in naturally contaminated and spiked samples of corn and peanut using the Aflatoxin Plate Kit. This kit is an enzyme-linked immunosorbent assay (ELISA) suitable for rapid testing of grains and peanuts. The assay was evaluated for ruggedness and linearity of the standard curve. The test kit results were then statistically evaluated for accuracy, precision, and correlation to a validated HPLC method (AOAC 994.08). The results were verified by an independent laboratory.

Sign in / Sign up

Export Citation Format

Share Document