The multiplexing technology that Eve Technologies uses is based on color-coded polystyrene beads. Bead coloration is achieved by utilizing different concentrations of red and infrared fluorophore dyes to create 100 uniquely-colored bead sets. Considering that these bead sets contain a unique color/fluorophore signature (that can be individually identified by the bead analyzer such as a Bio-Plex200), they can then be combined within the same assay well. For example, a capture antibody for IL-6 is coupled to a population of dark red beads, and a capture antibody for GM-CSF is coupled to a population of pink beads. These two antibody-coupled beads can now be mixed together forming a 2-plex. Each analyte is distinguished from the other because they are bound to differently colored/fluorescent beads.
The bead analyzer includes a dual-laser system and a flow-cytometry system. One laser activates the fluorescent dye within the beads which identifies the specific analyte. The second laser excites the fluorescent conjugate (streptavidin-phycoerythrin) that has been bound to the beads during the assay. The amount of the conjugate detected by the analyzer is in direct proportion to the amount of the target analyte. The results are quantified according to a standard curve.