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Faculty of Medicine, Comenius University in Bratislava
Comenius University in Bratislava
MEFANET (MEdical FAculties Educational NETwork) Project

RNDr. Vladimíra Ďurmanová, PhD., vladimira.durmanova(at)fmed.uniba.sk

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Serological methods II

Serological methods II

Serological methods are basic diagnostic methods used to identify antibodies and antigens in patient sample. Up-to-date serological methods involve detection of unknown concentration of antigen or antibodies in patient sample using specific labelled antibodies. In immunodiagnostics, the most used serological methods are: immunofluorescence (IF), enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA) and immunoblotting (Western blotting). Immunofluorescence (IF) detects antigens or antibodies by fluorochrome-labelled antibodies under fluorescence microscope. In the immunodiagnostics the IF is applied to detect autoantibodies in the tissue or patient serum. ELISA detects antigens and antibodies using enzyme-labelled antibodies following evaluation by spectrophotometry. In the immunodiagnostics, ELISA is widely used in quantitative analysis of autoantibodies, antibodies to vaccination antigens, bacterial and viral antigens, cytokines, etc. Radioimmunoassay (RIA) is a quantitative and sensitive diagnostic method to detect antigens or antibodies by radioisotope-labelled antibodies. In the immunodiagnostics, the RIA is used to detect the level of IgE involved in allergic reactions. Immunoblotting is a qualitative method based on protein separation in gel following by their transfer from the gel to the membrane and protein detection by enzyme-labelled antibodies. ELISA and immunoblotting belongs to the serological methods used in the confirmation of HIV positivity.

 
author: Vladimíra Ďurmanová | discipline: Immunology, Allergology | viewed: 2063x | published on: 1.10.2013 | last modified on: 1.10.2013

Serological methods I

Serological methods I

Serological methods are basic diagnostic methods used to identify antibodies and antigens in patient sample (serum, plasma). Agglutination and precipitation belong to classical serological methods that are used in diagnosis of infectious diseases (antibodies screening), serotyping of microorganisms and human blood group typing. Agglutination is based on reaction of particular (insoluble) antigen with antibodies, whereas precipitation involves reaction of colloidal (soluble) antigen and antibodies. In the immunodiagnostics an agglutination method as Coombs test is used to determine anti Rh antibodies in pregnant women (diagnosis of hemolytic disease of newborn). Another agglutination method involves latex agglutination that is used to determine, i.e. rheumatoid factor (diagnosis of autoimmune diseases) and hemagglutination method that can be used in, i.e. antibodies screening against Treponema pallidum, the main cause of syphilis. Radial immunodiffusion assay belongs to the precipitation method. This quantitative assay is based on immunodiffusion of antigens and antibodies in the gel and can be used in level analysis of complement components (C3, C4), inflammation proteins (CRP) and immunoglobulins (IgG, IgM and IgA). Modern precipitation methods performed in fluid involve nephelometry and turbidimetry.

 
author: Vladimíra Ďurmanová | discipline: Immunology, Allergology | viewed: 5946x | published on: 23.5.2013 | last modified on: 23.5.2013