Pesticide immunoassays specificity

Actual and perceived food safety concerns have necessitated an increase in the monitoring of foods for such natural contaminants as aflatoxins and for residues of pesticides. Immunoassays can provide rapid, simple, and relatively inexpensive methods for the detection of analytes with specificity and with sensitivities directed at the levels of concern. Particularly for aflatoxins, they are rapidly assuming a significant role in the monitoring of foods. However, the misuse of these techniques can potentially compromise any food safety improvement that may result from increased surveillance. Experiences of the Division of Contaminants Chemistry in the development, validation, and applications of immunoassays for natural toxins is discussed. 

Biopolymers are employed in many immunological techniques, including the analysis of food, clinical samples, pesticides, and in other areas of analytical chemistry. Immunoassays (qv) are specific, sensitive, relatively easy to perform, and usually inexpensive. For repetitive analyses, immunoassays compare very favorably with many conventional methods in terms of both sensitivity and limits of detection. 

In the following discussion, the detection of pesticides and veterinary dmgs in food animals by immunoassay will be described. Discussion will be organized by compound class, the specific analyte, and, finally, the tissues examined. The general principles described in the first part of this review provide the rationale in the applications described in the following pages. 

Cell components or metabolites capable of recognizing individual and specific molecules can be used as the sensory elements in molecular sensors [11]. The sensors may be enzymes, sequences of nucleic acids (RNA or DNA), antibodies, polysaccharides, or other reporter molecules. Antibodies, specific for a microorganism used in the biotreatment, can be coupled to fluorochromes to increase sensitivity of detection. Such antibodies are useful in monitoring the fate of bacteria released into the environment for the treatment of a polluted site. Fluorescent or enzyme-linked immunoassays have been derived and can be used for a variety of contaminants, including pesticides and chlorinated polycyclic hydrocarbons. Enzymes specific for pollutants and attached to matrices detecting interactions between enzyme and pollutant are used in online biosensors of water and gas biotreatment [20,21]. 

As part of SW-846, the EPA has validated and approved many immunoassay and colorimetric screening methods for a wide range of contaminants, such as petroleum fuels, pesticides, herbicides, PCBs, and explosives. Immunoassay technology uses the property of antibodies to bind to specific classes of environmental pollutants allowing fast and sensitive semiquantitative or qualitative detection. Colorimetric kits are based on the use of chemical reactions that indicate the presence of target analytes by a change in color. Table 3.9 presents a summary of EPA-approved screening methods and their detection capabilities. 

Applicability. Parker (4) points out that one can assume that workable radioimmunoassays can be developed "with all except the smallest or most unstable molecules." Once a useful antibody titer is obtained, often only very small changes in a generalized procedure are needed to obtain a workable assay. Although immunoassays would appear to be generally applicable to pesticide analytical problems they may be most useful in solving specific problems which appear intractable when classical procedures are used. Immunoassays are often most sensitive and specific when... 

Antibodies have been raised against representative compounds from the major classes of pesticides. Although the ELISA will be useful for individual analysis of a wide variety of compounds, if one needed to analyze several different compounds simultaneously in one matrix immunoassay may not be the method of choice, due to the large amount of controls and standards needed. However, it could be successfully used for the rapid screening of a large number of samples for the presence of specific types of pesticides and for confirmatory tests (Ji). The work reported here with paraquat,... 

Immunoassays offer much potential for rapid screening and quantitative analysis of pesticides in food and environmental samples. However, despite this potential, the field is still dominated by conventional analytical approaches based upon chromatographic and spectrometric methods. We examine some technical barriers to more widespread adoption and utilization of immunoassays, including method development time, amount of information delivered and inexplicable sources of error. Examples are provided for paraquat in relation to exposure assessment in farmworkers and food residue analyses molinate in relation to low-level detection in surface waters and bentazon in relation to specificity and sensitivity requirements built in to the immunizing antigen. A comparison of enzyme-linked immunosorbent assay (ELISA) results with those obtained from conventional methods will illustrate technical implementation barriers and suggest ways to overcome them. 

Applications of immunoassay to pesticide chemistry have been described which address some difficult problems in analysis by classical methods. These include stereospecific analysis of optically active compounds such as pyrethroids (38), analysis of protein toxins from Bacillus thuringiensis (5,37), and compounds difficult to analyze by existing methods, such as diflubenzuron (35) and maleic hydrazide (15 also Harrison, R.O. Brimfield, A.A. Hunter, K.W.,Jr. Nelson, J.O. J. Agric. Food Chem. submitted). An example of the excellent specificity possible is seen in assays for parathion (10) and its active form paraoxon (3). Some immunoassays can be used directly for analysis without extensive sample extraction or cleanup, dramatically reducing the work needed in typical residue analysis. An example of this is given in Figures 2 and 3, comparing the direct ELISA analysis of molinate in rice paddy water to the extraction required before GC analysis. 

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