Pesticides immunochemical methods

S.J. Gee, A.D. Lucas, and B.D. Hammock, Using immunochemical methods to analyze for biomarkers of exposure, in Methods of Pesticide Exposure Assessment, ed. PB. Curry,... 

LC-MS at present is only complementary to GC-MS. The LC-MS technique is applied for confirmations of positive results obtained by different immunochemical methods and for confirmation of the intake of drag(s). Nevertheless, the number of studies in which procedures for screening a single group of drugs, for example, benzodiazepines, amphetamines, fentanyl analogs (Fig. 16.1), and pesticides, or a subject... 

Aston, J.P., D. Britton, M. Wraith, and A. Wright (1992). Immunochemical methods for pesticide residue analysis. In T. Cairns and J. 

Several qualitative and quantitative immunochemical methods and their application to the analysis of environmental samples have been described for OP insecticides, a family that includes widely used pesticides such as azinphos-ethyl/methyl, dichlorvos, fenitrothion or fenthion, malathion, mevinphos, and parathion. Mercader and Montoya202 produced monoclonal antibodies against azinphos-methyl and developed an ELISA that was used for the analysis of water samples from different sources, reaching detectability levels near 0.05 pg I. Watanabe et al.203 reported the production of polyclonal antibodies and ELISA procedures to analyze fenitrothion in river, tap, and mineral water (LOD = 0.3 pg L ). Banks et al.204 produced polyclonal antibodies against dichlorvos, an organophosphate insecticide used for stored grain, which also cross-reacts with fenitrothion. Nishi et al.205 reported the first immunoassay for malathion. Residues of this insecticide have... 

The sensitivity of the overall analytical procedure depends upon many factors obviously including the type of sample to be analyzed and the skill of the analytical chemist. If an immunoassay is used to measure the amount of pesticide in a water sample by adding the water sample directly to the immunoassay, very high sensitivity may not be obtained although the assay will require very little time to perform. Alternatively, if the water sample is extracted and the immunoassay is employed only after several highly efficient cleanup steps, phenomenal sensitivity may be obtained at the expense of a large investment in time. In some situations, immunochemical methods may decrease the limit of detectability of a pesticide residue (77), but more importantly they may, in some cases, decrease the time and cost needed to reach a level of detectability as has been demonstrated with parathion (31). 

Possible Contributions of Immunochemical Methods to Pesticide Analysis. As Ercegovich (3 ) pointed out, it is unlikely that immunochemical methods will replace current, established analytical methods of pesticide analysis. However, the analytical chemist who carefully compares the attributes and deficiencies of immunochemical methods of analysis with other procedures is likely to find applications for which immunochemical methods offer distinct advantages. 

It is probable that in certain situations immunochemical methods will provide distinct advantages over conventional analytical methods. However, it is unlikely that immunochemical methods will completely replace current, established analytical methods of pesticide analysis (5.). This is in spite of the fact that chemical classes currently assayed by immunochemical techniques in clinical analytical labs contain the same type of functional groups as many pesticides. 

The adaptation of immunochemical methods for many classes of small compounds has made it possible to develop assays for pesticide residue detection. A cELISA was developed for methoprene, an analog of insect juvenile hormone. Because of its size, methoprene does not elicit an immune response by itself. However, by conjugating methoprene to a carrier protein it was made immunogenic in animals. A four-carbon spacer group was incorporated between methoprene and the carrier protein. The spacer was first coupled to methoprene acid by a series of protection/deprotection steps. 

Immunochemical methods for low molecular weight (<1000 D) environmental compounds, such as pesticides, nitro aromatics, and PAHs (polycyclic aromatic hydrocarbons) have been developed since more than 25 years (e.g. Hammock and Mumma, 1980 Hammock et al., 1990 Dankwardt, 2000). The most common method is the immunoassay, which is either carried out with (heterogeneous) or without (homogenous) separation steps. Heterogeneous formats that use microtitre plates, plastic tubes and/or membranes as solid supports are more commonly used. 

Immunochemical methods are rapidly gaining acceptance as analytical techniques for pesticide residue analysis. Unlike most quantitative methods for measuring pesticides, they are simple, rapid, precise, cost effective, and adaptable to laboratory or field situations. The technique centers around the development of an antibody for the pesticide or environmental contaminant of interest. The work hinges on the synthesis of a hapten which contains the functional groups necessary for recognition by the antibody. Once this aspect is complete, immunochemical detection methods may take many forms. The enzyme-linked immunosorbent assay (ELISA) is one form that has been found useful in residue applications. This technique will be illustrated by examples from this laboratory, particularly molinate, a thiocarbamate herbicide used in rice culture. Immunoassay development will be traced from hapten synthesis to validation and field testing of the final assay. 

Immunochemical methods have also been applied to the detection of bound pesticide residues in soil. These are formed by binding of pesticides to the organic matter of the soil, mainly humic and fulvic acids, and cannot be analyzed using common extraction and assay methods. Hahn et al. used Fab fragments labeled with a fluorescent dye to detect nonextractable residues of atrazine in soil from corn fields. The fluorescence signal obtained was related to the amount of bound atrazine in native soil samples determined by GC after supercritical methanol extraction. A noncompetitive sandwich lA for the analysis of bound residues based on HA-Ab and triazine Ab was developed by Ulrich et HA was extracted from soil, bound to the plates by the HA-Ab and the nonextractable triazine residues were detected by... 

M.J. Wraith, D.W. Britton, Immunochemical Methods for Pesticide Residue Analysis , Brighton Crop Protection Conference, British Crop Protection Council, London, 131-137,1988. 

Wraith, M. J. Britton, D. W. Immunochemical methods for pesticide residue analysis. Brighton CropProL Conf.-Pests IMs., (1), 131-7. 1988. 

Recent trends in pesticide analysis in food aims for reduced sample pretreatments or simplified methodologies (as QuEChERS approaches), the use of online purification processes, the use of new adsorbents (such as molecular imprinted polymers (MIPs) and nanomaterials) for the extraction and clean-up processes, and focused on the development of large multiresidue methods, most of them based on LC-MS/ MS. In spite of the relevant role of LC-MS/MS, GC-MS-based methods still play an important role in pesticide analysis in food. Despite the development achieved in the immunochemical approaches, the need for multi-residue methods has supported the development and use of instrumental techniques. 

The enzyme-linked Immunosorbent assay (ELISA) is a rapid Immunochemical procedure which can be used for trace analysis. We have applied the procedure to paraquat and other compounds difficult to analyze by the more classical methods. The Immunoassay for paraquat shows the practicality of the method for fortified and actual residue samples, and Is being compared with a gas chromatography procedure. Our work with the ELISA Illustrates that the Immunochemical technology can be used to solve problems encountered In pesticide residue analysis. 

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