Evaluation of Secreted Cytokines by Multiplex Bead-Based Assay (X MAP Technology, Luminex)

The Luminex XMAP technology permits the simultaneous evaluation of numerous cytokines in several types of biological fluids (plasma, serum, liquor, follicular fluids, etc.) and in cell supernatants. Thus, multiplexing allows to achieve a time/cost economy and ensures that all the measurements are performed in the same conditions. Simultaneous measurement of cytokines with a multiplex bead-based assay has some similarities with ELISA, in particular the use of anti-cytokine antibodies, but shows an important difference, the use of magnetic fluorescent beads coupled to anti-cytokine monoclonal antibodies. The magnetic microspheres (dyed internally with two florescent dyes) coupled with anti-cytokine monoclonal antibodies are incubated with samples and standards; after washing, the samples/standards are incubated with biotinylated anti-cytokine monoclonal antibodies; and finally, after other washings, with streptavidin-phycoerythrin solution. Luminex instrument identifies the different cytokines present in each well and converts the mean fluorescence intensity (MFI) of each measured cytokine in pg/ml, thanks to the software and the standard curves. This technique is applicable in basic and clinical research.