Development of an enzyme-linked immunosorbent assay for imazaquin herbicide

Monoclonal antibodies were developed that bind sarafloxacin, a fluoroquinolone approved by the Food and Drug Administration for use against Escherichia coli in poultry. Splenocytes from mice immunized with a bovine serum albumin−sarafloxacin conjugate were fused with SP2/0 myeloma cells, and hybridomas secreting antibodies against sarafloxacin were selected and cloned. An enzyme-linked immunoassay was developed, and 50% inhibition of control values ranged from 7.3 to 48.3 ppb using sarafloxacin as the competitor. Tissue samples were spiked with sarafloxacin, and the average percent recoveries at 10, 50, and 100 ppb were 132, 78, and 81%, respectively. Monoclonal antibodies exhibiting high relative affinity for sarafloxacin were also characterized for their ability to detect five structurally related quinolones. The specificity and cross-reactivities of these antibodies are discussed in relation to three-dimensional, computer-generated molecular models of the fluoroquinolones. Keywords: Sarafloxacin; fluoroquinolone; ELISA; immunoassay

Section: Introductionmentioning

confidence: 99%

Production and Characterization of Monoclonal Antibodies against Sarafloxacin and Cross-Reactivity Studies of Related Fluoroquinolones

Holtzapple

Buckley

Stanker

1997

“…Assay format appears to have an effect on the ability of an immunoassay to overcome these matrix effects. For analysis of environmental samples, competitive ELISAs that use immobilized antibody and an enzyme conjugate (Rubio et al, 1991;Schneider and Hammock, 1992;Wong and Ahmed, 1992;Lawruk et al, 1993a) perform better than competitive ELISAs that use free antibody in solution and a coating conjugate (Bushway et al, 1988;Feng et al, 1990;Goh et al, 1990). The purpose of this paper is to describe the development of competitive ELISAs for the herbicides fluroxypyr and triclopyr and outline the application of these ELISAs for use in quantitating concentrations of fluroxypyr and triclopyr in water and soil.…”

Section: Introductionmentioning

confidence: 99%

Fluroxypyr- and Triclopyr-Specific Enzyme-Linked Immunosorbent Assays: Development and Quantitation in Soil and Water

and

Hall

1996

Enzyme-linked immunosorbent assays (ELISAs) were developed to quantitate fluroxypyr [[(4-amino-3,5-dichloro-6-fluoro-2-pyridinyl)oxy]acetic acid] and triclopyr [[(3,5,6-trichloro-2-pyridinyl)oxy]acetic acid] in soil and water. The linear working range of the fluroxypyr ELISA was from 0.1 to 10 ng‚mL -1 with a limit of detection (LOD) of 0.1 ng‚mL -1 and an IC 50 value of 1.6 ng‚mL -1 . The linear working range of the triclopyr ELISA was from 0.1 to 5 ng‚mL -1 with a LOD of 0.1 ng‚mL -1 and an IC 50 value of 1.7 ng‚mL -1 . Cross-reactivity to selected pyridine metabolites and agrochemicals was determined. Significant cross-reactivity (within the linear working range of the ELISA) using the fluroxypyr ELISA was found only to the metabolite 4-amino-3,5-dichloro-6-fluoro-2-methoxypyridine. Significant cross-reactivity using the triclopyr ELISA was found only to the auxinic herbicide 2,4,5trichlorophenoxyacetic acid (2,4,5-T). The ELISAs accurately estimated fluroxypyr and triclopyr in water at concentrations as low as 0.1 ng‚mL -1 . Analysis of two different soil types with different textures (clay loam and sandy clay loam) required cleanup procedures using filtration and solid phase extraction to accurately estimate fluroxypyr and triclopyr concentrations.

“…Immunoassay technology is being demonstrated as a viable alternative for and a complement to traditional analytical methods for monitoring agrochemicals (Brecht et al, 1995;Schwalbe et al, 1984;Thurman et al, 1990;Wong and Ahmed, 1992). Most assays have followed the enzyme-linked immunosorbent assay (ELISA) format, in which measurements are made of the color produced from a colorless substrate by the action of an enzyme conjugated to either an antibody or an analyte molecule.…”

Section: Introductionmentioning

confidence: 99%

Determination of Imazethapyr Using Capillary Column Flow Injection Liposome Immunoanalysis

Lee

Durst

1996

A sensitive immunoanalysis system was developed for the quantitation of imazethapyr, the active ingredient in PURSUIT herbicide. Imazethapyr [5-ethyl-2-(4-isopropyl-4-methyl-5-oxo-2-imidazolin-2-yl)nicotinic acid] is one of the imidazolinone class of herbicides. The assay was based on sequential competitive binding of imazethapyr and liposomes for a limited number of antibody binding sites. A capillary tube (20 cm × 0.53 mm i.d.) with immobilized antibody was used as the immunoreactor column. Liposomes that entrap fluorescent molecules as the detectable label provide instantaneous, rather than time-dependent, enhancement, common with enzyme immunoassays. In this study, liposomes encapsulated carboxyfluorescein dye and were made antigenic by incorporating in the bilayer a phospholipid that had the analyte conjugated to its polar head group. The calibration curve for imazethapyr in Tris-buffered saline solution had a working range of 0.1−100 ng/mL. In the range between 1 and 100 ng/mL, recoveries from fortified tap and pond water samples ranged from 93 to 114%. Filtration was the only step needed for sample cleanup, and an assay could be performed in <10 min. Keywords: Imazethapyr; FILIA (flow injection liposome immunoanalysis); capillary immunocolumn; immunoassay; herbicide; liposomes