Enzyme immuno-assay

Viral structural proteins expressed by baculovirus infected insect cells assemble into multimeric structures that resemble viral core-like particles and virus-like particles (CLPs and VLPs, respectively). This presentation has brought the attention of researchers for the potential use of these structures as safe immunological reagents for virus or antibody detection in enzyme immuno assays, as vaccines, and more recently, as gene delivering systems for gene therapy [9]. 

We have measured FSH in unextracted urine on an AxSYM random-access immunoassay analyzer (Abbott laboratories, Abbott Park, IL) with a MEIA (microparticle enzyme immuno assay) reagent kit. In order to correct for dilution, creatinine was measured, and the urinary FSH was normalized for creatinine concentration. Urine and serum samples were obtained from 40 women between 32 and 55 years of age. All women were healthy, except for a benign gynecological illness for which they were admitted to our hospital. All women had normal renal function. On the day of operation, we took six serum samples from each patient, each at least an hour apart, in order to calculate the mean serum FSH concentration. During the same day, we collected an early-morning urine sample, 24-h urine sample, and a random void urine sample. 

High Performance Liquid Chromatography 7.10. Enzyme Immuno Assay... 

Maertlbauer et al. [34] developed a highly specific and sensitive enzyme immuno assay for the quantitative determination of natamycin in cheese rind. Rabbits were immunized with a natamycin-HSA conjugate to produce a specific antiserum. A labelled ligand was produced by coupling natamycin to horseradish peroxidase. The range of this ELISA test was between 0.2 and 2 ng per ml of sample solution. This allowed the determination of natamycin in cheese rind down to concentrations of 5 ng/dm2 or 0.1 mg/kg. The recovery in a range of 1 to 80 mg/kg was 76 to 84%. The cross-reactivity with amphotericin B and nystatin was <0.001%. 

Tjjssen, P. (1985). Practice and Theory of Enzyme Immuno assay, Elsevier, Amsterdam... 

Conjugated steroids were hydrolysed, extracted and measured with two enzyme immuno assays (EIA) specific for total estrogens and pregnanediol, respectively. Determinations of hormone concentrations were carried out as described by Meyer et al. (1997). 

Use of a surrogate end point that is quick and easy to obtain Permeation experiments using a radiolabeled, fluorescent, HPLC-detectable, or radio immuno assay/enzyme linked immuno sorbent assay-detectable marker necessitate the need of extensive sample handling and sample analysis. This accentuates the cost of sample analysis and overall time spent in characterizing the efficacy of formulations. Furthermore, current state of the art fluidics systems put a fundamental limit on the number of samples handled in a given time. 

Table 4 Examples of enzyme immuno-like assays...

Surugiu et al. [67] have introduced an Enzyme Immuno-Like Assays (EzILA) for the herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The label was a 2,4-D conjugate with the tobacco peroxidase (TOP) enzyme, which allows for both colorimetric and chemiluminescent detection. In this case, the polymer imprinted with 2,4-D was synthesized in the form of microspheres. In contrast, despite its higher binding capacity for radiolabeled 2,4-D, a conventional MIP prepared by bulk polymerization showed only weak binding of the 2,4-D-TOP tracer. 

One prominent use of organometallic complexes is in metallo-immuno assays. The traditional radio-linked immuno assay (RIA) is highly sensitive but has obvious disadvantages related to the use of radioactivity. Modern alternatives use colorimetric, fluorescence, or enzyme-linked detection schemes (ELISA). The idea of using non-radioactive metals for specific, highly sensitive detection in immuno assays was first mentioned by Gais, who used steroid... 

Fig. 1. (left) Identification of the immunoprecipitated enzyme after gel electrophoresis and autoradiography. This experiment was designed to test die specificity of the immuno-assay using either the free or immobilized antibody. Lane 1 shows total P S]-labelled enzyme obtained using the immobilized antibody lane 2, unmodified enzyme obtained after boronate chromatography lane 3, identical to lane 1 except that free antibody was used and lane 4, pre-immune serum control for lane 3. 

A bridge between natural and artificial macromolecular metal complexes is the interaction of metal ions/complexes with peptides/proteins [70], nucleic acids/DNA [71,72], enzymes [73], steroids [74], carbohydrates [75]. Biometal-organic chemistry concentrates on such complexes [15], The reason for the increasing interest in this field lays in medical applications of metal complexes [16,76] (cancer, photodynamic therapy of cancer -immuno-assays, fluorescence markers, enantioselective catalysis, template orientated synthesis of peptides) as exemplarily shown below. 

Doi, H., Shibata, H., Shoji, M., Sakai, S., and Akiyama, H. (2008). A reliable enzyme linked immuno-sorbent assay for the determination of walnut proteins in processed foods. /. Agric. Food Chem. 56, 7625-7630. 

King, D.P. et al., The use of monoclonal antibodies specific for seal immunoglobulins in an enzyme-linked immunosorbent assay to detect canine distemper virus-specific immunoglobulin in seal plasma samples, J. Immuno. Methods, 160, 163, 1993. 

Immunoaffinity chromatography (IAC), 6 400—402 12 137, 145 Immunoanalyzers, automated, 14 150 Immunoassay(s), 14 135-159. See also Immunoassay- DNA probe hybrid assays Immunoassay methods Immuno(bio)sensors antibody-antigen reaction, 14 136-138 basic technology in, 14 138-140 chemiluminescent, 14 150-151 classification of, 14 140-153 design of, 14 139-140 enzyme, 14 143-148 fluorescence, 14 148-150 highly specific, 14 153 historical perspective on, 14 136 microarrays and, 14 156—157 microfluidics in, 26 968—969 monoclonal versus polyclonal antibodies in, 14 152-153... 

Enzyme-linked immunosorbent assay (ELISA) is comparable to the immuno-radiometric assay except that an enzyme tag is attached to the antibody instead of a radioactive label. ELISAs have the advantage of nonradioactive materials and produce an end product that can be assessed with a spectrophotometer. The molecule of interest is bound to the enzyme-labeled antibody, and the excess antibody is removed for immunoradiometric assays. After excess antibody has been removed or the second antibody containing the enzyme has been added (two-site assay), the substrate and cofactors necessary are added in order to visualize and record enzyme activity. The level of molecule of interest present is directly related to the level of enzymatic activity. The sensitivity of the ELISAs can be enhanced by increasing the incubation time for producing substrate. 

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