Clinical immunoassays

ImmunO lSS iy. Chemiluminescence compounds (eg, acridinium esters and sulfonamides, isoluminol), luciferases (eg, firefly, marine bacterial, Benilla and Varela luciferase), photoproteins (eg, aequorin, Benilld), and components of bioluminescence reactions have been tested as replacements for radioactive labels in both competitive and sandwich-type immunoassays. Acridinium ester labels are used extensively in routine clinical immunoassay analysis designed to detect a wide range of hormones, cancer markers, specific antibodies, specific proteins, and therapeutic dmgs. An acridinium ester label produces a flash of light when it reacts with an alkaline solution of hydrogen peroxide. The detection limit for the label is 0.5 amol. 

The development of sensitive and inexpensive immunoassays for low molecular weight pesticides has been an important trend in environmental and analytical sciences during the past two decades. 0.27-29 jq design an immunoassay for a pesticide, one can rely on the immunoassay literature for agrochemicals, " but many of the innovations in clinical immunoanalysis are also directly applicable to environmental analysis. - Conversely, the exquisite sensitivity required and difficult matrices present for many environmental immunoassay applications have forced the development of technologies that are also useful in clinical immunoassay applications. In the following discussion we will describe widely accepted procedures for the development of pesticide immunoassays. 

Portable inununoassay kits have been developed for a limited number of Toxins. The kits are capable of identifying Toxins within 30 minutes. Kits for additional Toxins are imder development. Clinical immunoassays (ELISA), as well as chemical analytical methods for detection and identification of Toxins are available. These methods can take from 2 to 4 hours to identify individual Toxins even under ideal conditions. 

Clinical immunoassays (ELISA) are available as well as tradihonal techniques to culture and idenhfy the pathogen. 

Assay application. At this point major differences appear between the historical use of clinical immunoassays and the potential applications of environmental and pesticide immunoassays. Most clinical assays have been applied to simple or well defined and consistent matrices such as urine or serum. In contrast, most matrices likely to be analyzed for pesticides are more complex, less well defined, and more variable. The potential for serious problems with matrix effects in the environmental field is far greater than most clinical immunoassays have encountered. The application of immunoassays to environmental analysis requires sampling strategies, cleanup procedures, and data handling fundamentally similar to those presently in use in any good analytical lab. The critical factor in the success of immunochemical technology will likely be competence... 

Garrett, P.E., and Krouwer, J.S. (1985) Method evaluation II precision and sensitivity considerations. Journal of Clinical Immunoassay, 8, 165 168. 

Miller, J. J., and R. Valdes. 1992. Methods for calculating cross-reactivity in immunoassay. Journal of Clinical Immunoassay 15 97-107. 

Valdes, R., and J. J. Miller. 1992. Increasing the specificity of immunoassays. Journal of Clinical Immunoassay 15 87-96. 

R.P.C. Rodgers, Data Analysis and Duality Control of Assays A Practical Primer , in Clinical Immunoassay The State of the Art, ed. W.R. Butt, Marcel Dekker, New York, 253-308,1984. 

Immunoassays presently form probably the most active area of exploitation of CL, ECL and BL, as evidenced by the extensive commerciahzation of luminescent immunoassay analyzers and test kits. Among these, acridinium ester-based CL assays for alkaline phosphatase and amplified CL assays using peroxidase labels have become the most widespread [207, 208]. Besides immunosassays CL and BL have been used in nucleic acid assays [209-211] and in cellular studies concerning, for instance, phagocytosis [212]. ECL immunoassays based on ruthenium trisbipyridyl labels are now becoming popular in clinical immunoassay analyzers [213]. 

Anisotropy measurements are widely used in biochemistry and are even used for clinical immunoassays. One reason for this use is the ease with which these absolute values can be measured and compared between laboratories. 

See also Chemiluminescence Overview. Chiroptical Analysis. Derivatizatlon of Analytes. Enzymes Enzyme-Based Assays. Fluorescence Overview Ciinicai and Drug Appiications. Gas Chromatography Coiumn Technoiogy Mass Spectrometry. Immunoassays Overview. Immunoassays, Applications Clinical. Immunoassays, Techniques Enzyme Immunoassays Luminescence Immunoassays. Infrared Spectroscopy Overview. Liquid Chromatography Column Technology Normal... 

The immunometric-type assay has also been adapted for use with nonisotopic labels and is typically carried out in a heterogeneous format in which the antibody is immobilized on a solid support, such as a microtiter dish, membrane, or collection of beads. The canonical clinical immunoassay format in toady s laboratories is the enzyme-linked immunosorbent sandwich assay, which employs two antibodies, one to capture the analyte and the other to detect and quantify it. More details of the principles of these and other immunoassay techniques are given elsewhere in this encyclopedia. 

Most clinical immunoassays require neither prior extraction nor purification of the sample containing the analyte before they are added to the reaction containing the appropriate amoimts of high-affinity specific antibody and label. This is not the case, however, for analytes such as thyroxin or cortisol, which are tightly boimd to highly avid binding proteins that compete with the antibody for both the analyte and the label. Some of the more important protein-bound analytes are shown in Table 3. 

See also Immunoassays Overview. Immunoassays, Applications Clinical. Immunoassays, Techniques Radioimmunoassays Enzyme Immunoassays. Polymerase Chain Reaction. 

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