Luminex Assays
Luminex assays are microsphere (bead) based immunoassays capable of detecting multiple analytes within a single sample and a single test run. Luminex assays are utilized in a multitude of clinical laboratory and research applications that require high sensitivity and accuracy for detection of specific analytes, and together with multiplexing capabilities, provide rapid, high throughput, high accuracy, and versatility that optimizes sample bioanalysis.
How it Works
The principle of the Luminex assay:
- Dye coded beads coupled to analyte specific capture antibodies (or antigens if the analyte of study is an antibody) are mixed with a test sample. Antibody specific analytes are captured and bound to the beads.
- Secondary biotinylated antibodies specific for the analyte of interest form a complex containing the bead, analyte, and secondary biotinylated detection antibody. Phycoerythrin (PE), a fluorescent dye, conjugated to streptavidin is added and binds to the biotinylated secondary antibody. Alternatively, phycoerythrin is directly coupled to the secondary antibody thus eliminating potential background due to the natural affinity of biotin for a number of serum proteins.
- During the incubation of the sample (followed by a wash to remove unbound antibodies and other proteins) and the subsequent secondary antibody step, a labelled complex forms on the bead.
- After washing excess secondary antibody away, beads are read using a dual laser flow detection system or light emitting diode instrumentation. One laser distinguishes the spectral signal produced by individual beads that are each internally labelled with a unique fluorophore that produces a unique spectral signature. The second laser identifies and quantitates the second ‘signal’ produced by the PE fluorescence. As such, the assay allows for the simultaneous differentiation of individual bead populations by their spectral signature as well as identification and quantification of antibody bound to the bead.
This technology platform enables assay developers to conjugate distinct capture antibodies (or antigens) to specific bead populations with a distinct signature or fluorescent dye. Combining different bead populations enables the simultaneous multiplexing for the detection and quantification of multiple analytes from a single drop of sample. These features make the Luminex platform (and others that use similar principles) one of the most robust and cost-effective systems used in bioanalysis.