Quantitative analysis immunochemical assays

Assays for biological activity, where applicable, should be part of the pivotal stability studies. Appropriate physicochemical, biochemical and immunochemical methods for the analysis of the molecular entity and the quantitative detection of degradation products should also be part of the stability program whenever purity and molecular characteristics of the product permit use of these methodologies. 

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. 

Immunoreagents are well suited for quantitation of amplified DNA. However, the exponential nature of PCR amplification makes it difficult to extrapolate from the amount of amplified DNA to the amount of starting material. The most reliable quantitative PCR methods involve real-time assay of amplified DNA during the exponential phase of the amplification reaction. Because the analyte of interest is usually the DNA template, not the amplification product, immunochemical methods at their present stage of development are generally best applied to an unamplified template where concentrations permit direct analysis or for qualitative assays. 

Immunochemical analysis is a convenient approach for environmental analysis provided appropriate antibodies and format have been developed. Often, the assay depends on a colorimetric determination for its quantitative result. The advantage of immunochemical analysis is its specificity as well as its speed. Specific references should be consulted on applications. 

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