Quantitation of Aflatoxin B1-N7-Guanine Adduct in Urine by Enzyme-Linked Immunosorbent Assay Coupled with Immunoaffinity Chromatography

Abstract IndiaA specific and sensitive method to quantitate aflatoxin B1-N7-guanine adduct in urine samples by immunoaffinity chromatography coupled with indirect competitive enzyme-linked immunosorbent assay (ELISA) is reported. A novel in vitro method to synthesize an antigen (bovine serum albumin-guanine-aflatoxin B1) and its use to produce polyclonal antibodies specific to the hapten aflatoxin B-N7-guanine are discussed. An indirect competitive ELISA developed to quantitate aflatoxin adduct showed a 50% inhibition at 15.6 pmol aflatoxin B1-N7-guanine (y=66.73+ (-19.8)x, r=-0.997). Interference by aflatoxin B1 was less than 5%, and no interference by guanine, aflatoxin Gι and aflatoxin M1 was observed. An immunoaffinity column was also developed, by using these polyclonal antibodies, for single-step purification of aflatoxin B1-N7-gua-nine. The immunoaffinity column and the indirect competitive ELISA were evaluated and validated by quantitation of aflatoxin B1-N7-guanine in urine from rats dosed with aflatoxin B1 (1 mg/kg body weight). Spiking studies with standard aflatoxin B1-N7-guanine adduct at 2 and 4 μg/mL phosphate- buffered saline gave 96 and 100% recoveries, respectively, for immunoaffinity cleanup column. The method also was tested successfully for quantitating aflatoxin B1-N7-guanine adduct in spiked human urine. Recoveries of the adduct were 79-90%.

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Simultaneous Determination of Aflatoxin B1 and Aflatoxin M1 in Food Matrices by Enzyme-Linked Immunosorbent Assay

Food Analytical Methods ◽

10.1007/s12161-012-9484-5 ◽

2012 ◽

Vol 6 (3) ◽

pp. 767-774 ◽

Cited By ~ 40

Author(s):

Wenxiao Jiang ◽

Zhanhui Wang ◽

Greta Nölke ◽

Jing Zhang ◽

Lanlan Niu ◽

...

Keyword(s):

Simultaneous Determination ◽

Immunosorbent Assay ◽

Aflatoxin B1 ◽

Aflatoxin M1 ◽

Enzyme Linked Immunosorbent Assay ◽

Food Matrices

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Application of ELISA to Retail Survey of Aflatoxin B1 in Peanut Butter

Journal of Food Protection ◽

10.4315/0362-028x-49.10.792 ◽

1986 ◽

Vol 49 (10) ◽

pp. 792-795 ◽

Cited By ~ 13

Author(s):

BHANU P. RAM ◽

L. PATRICK HART ◽

RICHARD J. COLE ◽

JAMES J. PESTKA

Keyword(s):

Immunosorbent Assay ◽

Aflatoxin B1 ◽

Coefficient Of Variation ◽

Simple Procedure ◽

Peanut Butter ◽

Competitive Elisa ◽

Routine Screening ◽

Enzyme Linked Immunosorbent Assay

A simple procedure was devised for the routine screening of aflatoxin B1 (AFB1) in peanut butter using enzyme-linked immunosorbent assay (ELISA). Peanut butter samples (5 g) were artificially contaminated with AFB1 and extracted by blending with 25 ml of 55% methanol and 10 ml of hexane. The extract was filtered and aqueous filtrate analyzed by a direct competitive ELISA. Recovery of AFB1 added to peanut butter samples ranged from 85 to 112%, with an average inter-well coefficient of variation of 18.4%. The inter-assay coefficient of variation was 22.7%. Using this procedure, only 3 of 63 commercial samples of peanut butter had detectable levels (>5.0 μg/kg) of AFB1.

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Development of an Immunoglobulin M (IgM) Capture Enzyme-Linked Immunosorbent Assay for Detection of Equine and Swine IgM Antibodies to Vesicular Stomatitis Virus

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.8.3.475-481.2001 ◽

2001 ◽

Vol 8 (3) ◽

pp. 475-481 ◽

Cited By ~ 10

Author(s):

En-min Zhou ◽

Jose Riva ◽

Alfonso Clavijo

Keyword(s):

Vesicular Stomatitis Virus ◽

Immunosorbent Assay ◽

Polyclonal Antibodies ◽

Competitive Elisa ◽

Immunoglobulin M ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Igm Antibodies ◽

Vesicular Stomatitis ◽

Elisa Plate

ABSTRACT An immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (MC-ELISA) was developed for the detection of primary infection of vesicular stomatitis virus (VSV) in equine and swine sera. The test was based on the use of biotinylated sheep antibodies against equine or swine IgM molecules bound to a streptavidin-coated ELISA plate. The captured IgM antibodies were detected by application of antigens prepared from the New Jersey and the Indiana VSV serotypes (VSV-NJ and VSV-IN, respectively) and mouse polyclonal antibodies against VSV-NJ and VSV-IN. The MC-ELISA was compared to a competitive ELISA (C-ELISA) and the standard microtiter serum neutralization (MTSN) assay by testing serum samples from horses and pigs experimentally infected with VSV-NJ or VSV-IN. The MC-ELISA detected specific homologous IgM antibodies from equine and swine sera as early as 5 and 4 days postinfection (DPI), respectively, and as late as 35 DPI. The MTSN test also detected antibodies as early as 5 DPI and as late as 160 DPI. In a similar fashion, the C-ELISA detected antibodies from 6 to 7 DPI and as late as 160 DPI. These results demonstrated that the MC-ELISA is a useful test for serodiagnosis of primary VSV infection in horses and pigs.

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The Development of a Quantitative ELISA for Aflatoxin B1 in Ground Peanuts Using Antibodies Isolated From the Yolk of a Laying Hen

Journal of Food Protection ◽

10.4315/0362-028x-57.11.1022 ◽

1994 ◽

Vol 57 (11) ◽

pp. 1022-1024 ◽

Cited By ~ 4

Author(s):

SUZHEN LI ◽

RONALD R. MARQUARDT ◽

ANDREW A. FROHLICH ◽

HAO XIAO ◽

JAMES R. CLARKE

Keyword(s):

Immunosorbent Assay ◽

Aflatoxin B1 ◽

Laying Hens ◽

Egg Yolk ◽

Laying Hen ◽

Competitive Elisa ◽

Enzyme Linked Immunosorbent Assay ◽

Coating Antigen ◽

Extraction Protocol

Antibodies directed against Aflatoxin B1 (AFB1) were produced in laying hens, isolated from egg yolk and applied in an enzyme-linked immunosorbent assay (ELISA) for AFB1 in ground peanuts. The ELISA sensitivity was improved by reduction of the amount of the coating antigen absorbed onto the microplate well surfaces. The main aflatoxins B1, B2, G1, G2, M1, aflatoxicol, sterigmatocystin were found to cross-react in the competitive ELISA, 100, 25, 23, 4, 0, 0, 0%, respectively. Aflatoxin B1 could be reproducibly detected and quantified in spiked ground peanuts at levels greater than 5 ppb using a hexane and methanol solvent based peanut extraction protocol.

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Aflatoxin M1 in Nepalese sera, quantified by combination of monoclonal antibody immunoaffinity chromatography and enzyme-linked immunosorbent assay

Carcinogenesis ◽

10.1093/carcin/14.6.1233 ◽

1993 ◽

Vol 14 (6) ◽

pp. 1233-1235 ◽

Cited By ~ 13

Author(s):

Hiroki Okumura ◽

Osamu Kawamura ◽

Seishi Kishimoto ◽

Akihiro Hasegawa ◽

Santosh M. Shrestha ◽

...

Keyword(s):

Monoclonal Antibody ◽

Immunosorbent Assay ◽

Immunoaffinity Chromatography ◽

Aflatoxin M1 ◽

Enzyme Linked Immunosorbent Assay

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Enzyme-Linked Immunosorbent Assay of Mycotoxins Using Nylon Bead and Terasaki Plate Solid Phases

Journal of Food Protection ◽

10.4315/0362-028x-47.4.305 ◽

1984 ◽

Vol 47 (4) ◽

pp. 305-308 ◽

Cited By ~ 16

Author(s):

J. J. PESTKA ◽

F. S. CHU

Keyword(s):

Immunosorbent Assay ◽

Aflatoxin B1 ◽

Linear Response ◽

Screening Method ◽

Detection Limits ◽

Microtiter Plate ◽

Aflatoxin M1 ◽

Enzyme Linked Immunosorbent Assay ◽

Solid Phases ◽

New Procedures

Nylon beads and Terasaki plates were tested as solid phases for the enzyme-linked immunosorbent assay (ELISA) of the mycotoxins aflatoxin B1 (AFB1), aflatoxin M1 (AFM1) and T-2 toxin. Both methods had detection limits comparable to that of mycotoxin microtiter plate ELISAs. Using the nylon bead ELISA, ELISA competition curves for AFB1, AFM1 and T-2 toxin exhibited linear response between 1.0 to 100, 0.1 to 100, and 0.1 to 10.0 ng/ml, respectively. Response ranges for Terasaki plate ELISAs of AFB1, AFM1 and T-2 toxin were 1.0 to 50, 0.05 to 0.50, and 0.5 to 1.0 ng/ml, respectively. The new procedures did not require specialized instrumentation and may be used as an economical screening method for mycotoxins in the field and to diagnose certain mycotoxicoses.

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Determination of Azadirachtin in Agricultural Matrixes and Commercial Formulations by Enzyme-Linked Immunosorbent Assay

Journal of AOAC International ◽

10.1093/jaoac/84.4.1001 ◽

2001 ◽

Vol 84 (4) ◽

pp. 1001-1010 ◽

Cited By ~ 8

Author(s):

Kalidindi Hemalatha ◽

Namburi B K Venugopal ◽

Beedu S Rao

Keyword(s):

Immunosorbent Assay ◽

Cross Reactivity ◽

Competitive Elisa ◽

Enzyme Linked Immunosorbent Assay ◽

Agricultural Commodities ◽

Indirect Competitive Elisa ◽

Capture Assay ◽

Ic50 Value ◽

Antibody Capture

Abstract An enzyme-linked immunosorbent assay (ELISA) was developed for azadirachtin (aza), a biopesticide from the neem tree (Azadirachta indica A. Juss). The immunogen was synthesized by epoxidation using the furan ring in the aza molecule. Rabbits were immunized with either bovine serum albumin (BSA)-azadirachtin or ovalbumin (OA)-azadirachtin conjugate. Evaluation of the antisera by antibody capture assay showed that the antibody titer of antisera raised against OA-aza was 1:30 000. An indirect competitive ELISA was developed with BSA-azadirachtin as coating antigen and aza-specific antibodies raised against OA-aza immunogen. The immunoassay showed an inhibitory concentration (IC50) value of 75 ppb, with a range of detection from 0.5 to 1000 ppb for azadirachtin [based on regression analysis, y = 85.87 (−18.89x); r2 = −0.97]. Cross-reactivity of the antibodies with 2 aza- derivatives (22,23-dihydro-23β-methoxy azadirachtin and 3-tigloylazadirachtol) was 33 and 29%, respectively. The indirect competitive ELISA was validated and evaluated by quantitating aza in spiked agricultural commodities and from neem formulations. Azadirachtin was spiked into 5 different agricultural commodities: tomato, brinjal, coffee, tea, and cotton seed at 500 and 1000 ppb and recovered at 62–100%. In samples drawn from 6 lots, the aza content in neem-seed kernels ranged from 0.1 to 0.15%; in commercial neem formulations the content ranged from 200 to 2000 ppm. The method developed may be applied to environmental monitoring of aza and quality assurance studies of aza-based commercial formulations.

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Indirect competitive ELISA and colloidal gold-based immunochromatographic strip for amantadine detection in animal-derived foods

Analytical Methods ◽

10.1039/c8ay02806k ◽

2019 ◽

Vol 11 (15) ◽

pp. 2027-2032

Author(s):

Mingfei Pan ◽

Jingying Yang ◽

Shijie Li ◽

Guozhu Wang ◽

Junping Wang ◽

...

Keyword(s):

Immunosorbent Assay ◽

Colloidal Gold ◽

Antiviral Drug ◽

Competitive Elisa ◽

Enzyme Linked Immunosorbent Assay ◽

Immunochromatographic Strip ◽

Indirect Competitive Elisa

In this research, an indirect competitive enzyme-linked immunosorbent assay (ELISA) and colloidal gold-based immunochromatographic (GICG) strip were developed for the detection of the antiviral drug amantadine (AM) in animal-derived foods.

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