LEGENDplex™ Multiplex Analysis
In cancer states, measuring soluble biomarkers is important in understanding immune states, tumor status, or tumor environments. BioLegend’s LEGENDplex™ bead-based immunoassays quantify multiple soluble analytes simultaneously in biological samples using a flow cytometer. LEGENDplex™ kits are provided with predefined panels, ranging from 3 to 13 specificities, or customers can mix and match any subset within each predefined panel using their Mix and Match system.
BioLegend’s LEGENDplex™ is a bead-based immunoassay that utilizes the same basic principles of sandwich immunoassays, whereby a soluble analyte is captured between two antibodies.
- Bead populations come in two sizes and differing levels of APC fluorescence, allowing them to be clearly distinguished from each other. Each bead set is conjugated with a specific antibody on the surface and serves as the capture bead for that particular analyte. When a selected panel of capture beads are mixed together and incubated with an unknown sample containing target analytes, each analyte will be bound by its specific capture bead.
- After washing, biotinylated detection antibodies are added and each detection antibody will bind to its specific analyte bound on the capture beads, thus forming capture bead-analyte-detection antibody sandwiches. Streptavidin-phycoerythrin (SA-PE) is subsequently added, which will bind to the biotinylated detection antibodies, providing fluorescent signal with intensities in proportion to the amount of bound analyte.
- For each bead population, the PE signal fluorescence intensity is then quantified using a flow cytometer.
- The concentration of a particular analyte is determined based on a known standard curve using the LEGENDplex™ data analysis software.