Chemiluminescent immunoassays (CLIA) combine the advantages of chemiluminescence sensitivity with the specificity of an immunoreaction. In this technique, an enzyme converts a substrate into a chemiluminescent signal that is emitted as photons of light in the visible or near visible range. The amount of light produced correlates to the amount of target analyte present in the sample.
CLIA assays provide a sensitive, high-throughput alternative to conventional colorimetric ELISA methods. Although the underlying principles are the same, the colorimetric substrate is replaced by a chemiluminescent substrate in CLIA assays.
In general, chemiluminescent substrates should provide the following advantages:
* Microplate selection: to develop a reliable CLIA assay, it is crucial to select a microplate that is suitable for luminescence. Plates are usually opaque white with solid white or clear-bottomed wells to avoid background and cross talk.
Ultimately, the target analyte is quantified by measuring the light intensity generated by the chemiluminescent immunoreaction. Luminescence is measured in Relative Light Units (RLU) by a luminometer, where RLUs are typically proportional to the amount of target analyte present in the sample.
Importantly, CLIA assays are one of the most sensitive detection methods due to signal multiplication and amplification. The technique can detect small concentrations of biological molecules (limit of detection = zeptomole 10-21 mol). In the presence of enhancers, chemiluminescent reactions occur for even longer without a reduction in the light output. Substrate can be added only minutes before detection and there is no requirement for a long incubation period nor a need for stopping reagents, as is the case with conventional colorimetric ELISA assays.
Find detailed protocols to perform direct and indirect CLIA and ELISA assays.
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Download our brochure for a comprehensive list of ELISA kits and to search ELISA analysis kits by target or gene name which can be customized based on your CLIA assay needs.
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