ANALYTICALTECHNIQUESASSIGNMENTELISA: ELISA (Enzyme-linked immunosorbent assay) is the testhas evolved from other types of immunoassays in the early 1970s and is now oneof the most advanced widely clinical, translational, used laboratory techniquein as well as clinical medicine. This assay isalso used for detecting and quantifying substances such as peptides, proteins,antibodies and hormones. ELISA test detects and measures antibodies in your blood. This test canbe used to determine if you have antibodies related to certain infectiousconditions.APPLICATIONS:· ELISA can be applied to determinationof serum antibody concentrations ina virustest.
· Applications of ELISAhave also been found in homepregnancy test, and in the foodindustry when detecting potential foodallergens such as milk, peanuts, walnuts, almonds, and eggs.· ELISA can also be used in toxicology as arapid presumptive screen for certain classes of drugs.· The ELISA was widelyused in various of areas such as Immunology, Biological Pharmacy, Diagnostic industry.· Detection of antibodiesin blood sample for past exposure to disease.Eg; lung diseases,trichinosis, HIV, and bird fluPRINCIPLE: Enzyme-linked Immunosorbent Assays(ELISAs) combine the specificity of antibodies with the sensitivity of simpleenzyme assays, by using antibodies or antigens coupled to an easily-assayedenzyme.
ELISAs can provide a useful measurement of antigen or antibodyconcentration. There are two main variations on this method: The ELISA can beused to detect the presence of antigens that are recognized by an antibody orit can be used to test for antibodies that recognize an antigen.· A sensitive immunoassay that uses an enzymelinked to an antibody or antigen as a marker for the detection of a specificprotein, especially an antibody or antigen.
· ELISAinvolves detection of “analyte” in a liquid sample using liquid reagent or drystrips.· In dry analysis, strip can be used inreflectrometry. The quantitative reading usually based on detection ofintensity of transmitted light by spectrophotometry of specific wavelength.
· The sensitivity of detection depends onamplification of signal during the analytic reaction. In some enzymaticreaction, the signal generated by enzyme which are liked to the detectionreagents in fixed proportions to allow accurate quantification.TYPES OF ELISA:•Direct ELISA•Indirect ELISA •Sandwich ELISA •Competitive ELISADirect ELISA:Itis suitable for the detection of proteinaceous antigens and may require pre-purificationof sample. It is performed when desired antibody is available in apre-conjugated state.Indirect ELISA: Theprimary antibody is not conjugated, then indirect ELISA is required in which aconjugated secondary antibody is targeted to the isotope of the primaryantibody.Sandwich ELISA It quantifies themeasure of antigen between two layers of antibodies.
Theantigen to be measured must contain no less than two antigenic locales equippedfor official to counter acting agent, since no less than two antibodies act inSandwich.Competitive ELISA: Inthis sort neutralizer is initially brooded in arrangement with a specimencontaining antigen. The Antigen-immunizer blend is then added to the microtitrewell which is covered with antigen.
The more the antigen show in the example,the less free immune response will be accessible to tie to the antigen-coveredwell. Subsequent to washing the well, compound conjugated auxiliary counteracting agent particular for isotype of the essential neutralizer is added todecide the measure of essential immune response bound to the well.