Enzyme-linked immuno sorbent assay

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. 

ELISA enzyme linked immuno sorbent assay... 

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. 

It is necessary to give some background information to understand the principle underlying immunochemical tests such as ELISA (Enzyme Linked Immuno Sorbent Assay). Such information is not always available to chemists, since chemistry and immunology have traditionally been separate disciplines. 

Fig. 1. Comparison of enzyme-linked immuno sorbent assay (ELISA, left) and immuno-polymerase chain reaction (IPCR, right). During ELISA, an antibody-enzyme conjugate is bound to the target antigen. The enzyme converts a substrate in solution to a detectable product. In IPCR, the antibody-enzyme conjugate is replaced by an antibody-DNA conjugate. The subsequent addition of a DNA polymerase enzyme (e.g., Taq), nucleotides and a specific primer pair uses the antibody-linked DNA marker sequence as a template for amplification of the DNA. The PCR product is finally detected as an indicator of the initial amount of antigen.

Fig. 3. Comparison of different enzyme-linked immuno sorbent assay (ELISA) methods adapted for immuno-polymerase chain reaction (IPCR). Dependent on the purification grade of the sample to be analyzed and the availability of specific and functionalized antibodies, several typical ELISA protocols were adapted to IPCR. In the direct approach (A), the pure antigen is immobilized to the microplate surface and subsequently detected by a labeled specific antibody. If no labeled antibody is available (e.g., because of unpurified ascites fluid containing the antibody or loss in activity following labeling), a standardized labeled secondary species-specific antibody is used for detection of the primary antigen-specific antibody (B). For the detection of the antigen from matrices such as serum, plasma, tissue homogenate, and so on, a capture antibody immobilized to the microplate surface was used either in a direct (C) or indirect (D) sandwich approach, with the latter one additionally including a secondary species-specific detection antibody. For different methods of coupling antibody and DNA, abbreviated by in this figure, compare Fig. 2. Note that protein A chimeras (Fig. 2A) are not compatible with capture antibodies (Fig. 3C, D).

Schlaeppi, J-M., W. Fory, and K. Ramsteiner (1989). Hydroxyatrazine and atrazine determination in soil and water by enzyme-linked immuno-sorbent assay using specific monoclonal antibodies. J. Agric. Food Chem., 37 1532-1538. 

ADA BEVS BHK CHO ELISA mAb MDCK MMR SCID tPA VLP adenosine deaminase deficiency baculovirus expression vector system baby hamster kidney cell line Chinese hamster ovary cell line enzyme linked immuno sorbent assay monoclonal antibodies Madin-Darby canine kidney epithelial cells measles, mumps, rubella severe combined immunodeficiency plasminogen activator virus-like particle... 

The basic tool for detecting antibodies binding to a specific antigen is the ELISA, or Enzyme Linked Immuno-Sorbent Assay [3]. In this assay, antigen-specific antibodies are detected by antigen-mediated attachment to a solid support and secondary detection with an Fc-specific enzyme-labeled secondary antibody (Fig. 2). For hapten immunization, ELISA is performed using a carrier protein, typically BSA (bovine serum albumin), different from the carrier protein used for immunization. In this manner only antibodies with binding specificity to the hapten are revealed by the assay. 

As E. coli 0157 H7 is the most implicated STEC serotype in human cases, numerous immuno-based methods for the detection of this specific serotype have been documented. They include conventional Enzyme Linked Immuno Sorbent Assays (ELISA) in microplates, one-step immuno-detection systems, fully automated systems and various non enzymatic immunological based systems like Rapid Plate Latex Agglutination (RPLA), Immuno-Magnetic Separation (IMS) or immuno-chromatography (Table 3). 

Energy dispersive spectroscopy Ethylene diamrnine tetraacetic acid Enzyme linked immuno sorbent assay Electron microscopy... 

Screening test kits - enzyme linked immuno-sorbent assay (ELISA)... 

Gasc6n J, Oubina A, Barcelb D. (1997). Detection of endocrine-disrupting pesticides by enzyme-linked immuno-sorbent assay (ELISA) Application to atrazine. Trends in Analytical Chemistry 16 554-562. 

Berger JA, Berger LR (1979) Development of a colorimetric enzyme linked immuno sorbent assay test to assay ciguatoxin in fish tissue. Rev Int Oceanogr Med 53-54(0) 23-32. 

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