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Autoimmune Diagnostics

Enzyme-linked Immunosorbent Assay - ELISA

Once many antigens were identified at the molecular level, more and more ELISA techniques were implemented based on the use of purified or recombinant antigens.

The ELISA test determines whether a specific protein, such as an autoantibody, is present in a serum sample. The appropriate antigen is first absorbed to the walls of the wells of a microtitre plate.

Pre-coated microtitre plate
© Orgentec Diagnostika

One microtitre plate can be used to carry out up to 96 simultaneous tests. Each microtitre plate is comprised of 12 individual strips with 8 wells each. These strips can be used individually. Commercial ELISA kits provide a ready-to use pre-coated microtitre plate.

Components of a commercially available ELISA kit
© Orgentec Diagnostika

ELISA kit: separable microtitre plate coated with antigen, A-F: standard solutions for calibration, 1 and 2: positive and negative controls, concentrates for wash- and sample buffers, conjugate solution (peroxidase-coupled anti-human-IgG), substrate and stop solutions.

ELISAs are more objective than IFT1), are easy to carry out, and are more readily standardized. Because of their high sensitivity, they sometimes have the disadvantage of detecting autoantibodies that react at low titre levels, which may lead to false positives. False negative results also occur, especially when recombinant antigens do not express all relevant epitopes.

A further advantage of ELISAs is that the technology has developed to the point that the instruments can carry the tests out in large numbers. This is particularly advantageous in large clinics, where the sample volume can be high.

In step-wise diagnosis, ELISAs often serve to confirm IFT screening tests and to differentiate antibodies detected by immunofluorescence.

1)IFT: immunofluorescence test
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