Enzyme-linked immunosorbent assay screenings

Garthwaite, L, Ross, K.M., Miles, C.O., Briggs, L.R., Towers, N.R., Borrell, T., and Busby, R, Integrated enzyme-linked immunosorbent assay screening system for amnesic, neurotoxic, diarrhetic, and paralytic shellfish poisoning toxins found in New Zealand. J. AOACInt., 84, 1643, 2001. 

Cole, R. J. Domer, J. W. Kirksey, J. W. Dowell, F. E. Comparison of visual, enzyme-linked immunosorbent assay screening, and HPLC methods in detecting aflatoxin in farmers stock peanut grade samples. Peanut Sci., 15 61-3. 1988. 

TR. Dombrowski, E.M. Thurman, and G.B. Mohrman, A first application of enzyme-linked immunosorbent assay for screening cyclodiene insecticides in ground water, in Environmental Immunochemical Methods, ed. J.M. Van Emon, C.L. Ger-lach, and J.C. Johnson, American Chemical Society, Washington, DC, pp. 148-154 (1996). 

The most commonly used screening method for HIV is an enzyme-linked immunosorbent assay, which detects antibodies against HIV-1 and is both highly sensitive and specific. False positives can occur in multiparous women in recent recipients of hepatitis B, HIV, influenza, or rabies vaccine following multiple blood transfusions and in those with liver disease or renal failure, or undergoing chronic hemodialysis. False negatives may occur if the patient is newly infected and the test is performed before antibody production is adequate. The minimum time to develop antibodies is 3 to 4 weeks from initial exposure. 

Most of the analytical methods for the analysis of pesticides in food are based on instrumental approaches based on chromatography coupled to mass spectrometry. However, a great effort of development has been paid to develop rapid screening methods based on biological methods, such as, enzyme linked immunosorbent assays (ELISA). 

Benzodiazepines are an important group of drugs with tranquilizing properties. Available immunochemical methods include radioimmunoassays (164, 165), a radioreceptor assay (166), and nonseparation immunoassays such as the widely used enzyme-monitored immunotest (EMIT) and fluorescent polarization immunoassays (167, 168). Such assays generally require sophisticated apparatus and dedicated laboratories. However, a relatively simple enzyme-linked immunosorbent assay was recently described for screening benzodiazepines in urine (169). 

An ELISA (enzyme-linked immunosorbent assay) allows for rapid screening and quantification of the presence of an antigen in a sample (Fig. 5-28b). Proteins in a sample are adsorbed to an inert surface, usually a 96-well polystyrene plate. The surface is washed with a solution of an inexpensive nonspecific protein (often casein from nonfat dry milk powder) to block proteins introduced in subsequent steps from also adsorbing to these surfaces. The surface is then treated with a solution containing the primary antibody—an antibody against the protein of interest. Unbound antibody is washed away and the surface is treated with a solution containing antibodies against the primary antibody. These secondary antibodies have been linked to an enzyme that catalyzes a reaction that forms a colored product. After unbound secondary antibody is washed away, the substrate of the antibody-linked enzyme is added. Product formation (monitored as color intensity) is proportional to the concentration of the protein of interest in the sample. 

Test wells exhibiting growth for the presence of monoclonal antibodies when the hybridoma growth covers approx 25% of the base of the cell well. The assay system used should mimic the final test format required. Commonly, Triple Antibody Sandwich (TAS) (5) enzyme-linked immunosorbent assay (ELISA) is use for screening hybridomas for specific antibody. It is important when testing hybri-... 

Situ, C., E. Gratters, P. van Wichen, et al. 2006. A collaborative trial to evaluate the performance of a multi-antibiotic enzyme-linked immunosorbent assay for screening five banned antimicrobial growth promoters in animal feeding stuffs. Anal. Chim. Acta 561 62-68. 

Lu, H., G. Conneely, M.A. Crowe, et al. 2006. Screening for testosterone, methyltestosterone, 19-nortes-tosterone residues and their metabolites in bovine urine with enzyme-linked immunosorbent assay (ELISA). Anal. Chim. Acta 570 116-123. 

Particle Concentration Fluorescence Immunoassay. The PCFIA is a solid-phase immunoassay in which proteins are attached to polystyrene particles by adsorption or covalent coupling for the solid phase and fluorescent-labeled reagents are utilized for product detection.22 The general principles of the assay are similar to those of the enzyme-linked immunosorbent assay (ELISA), which has been reviewed extensively elsewhere.23 PCFIAs are performed in specially designed 96-well format Fluoricon assay plates utilizing an automated Screen Machine (Idexx Corporation, Research Product Division, Portland, ME). 

Serum samples are analyzed for antibody titer. When the titer is sufficiently high, the animal is bled. Serum samples are generally screened for the presence of the antibody by enzyme-linked immunosorbent assay (ELISA). ELISA methods are discussed later in this chapter. 

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