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BioLegend LEGENDplex
BioLegend’s LEGENDplex™ bead-based immunoassays quantify multiple soluble analytes simultaneously in biological samples using a flow cytometer.
Principle of the Assay
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. 
  1. 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.
  2. 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.
  3. For each bead population, the PE signal fluorescence intensity is then quantified using a flow cytometer.
  4. The concentration of a particular analyte is determined based on a known standard curve using the LEGENDplex™ data analysis software.
BioLegend's 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 our Mix and Match system. If you would like to combine specificities across different predefined panels, i.e., Human Cytokine and Human Chemokine, you can make a request to our Custom Solutions Team or email CST@biolegend.com.

Each kit provides sufficient reagents for at least 100 tests. The assay can be performed in a 96-well filter plate, allowing you to run 40 samples plus 8 standard curve titrations in duplicate, or they can be performed in FACS tubes, where duplicate titrations may not be necessary.

For analysis of LEGENDplex™ assays, BioLegend provides the LEGENDplex™ software for free.  A dongle (USB drive) with a limited number of analysis runs is required in order to use the software. Dongles are provided with every kit ordered. To download the software, click on the Software tab above.

Fluorescent LEGENDplex™ Carboxyl Beads are also available for do-it yourself conjugations to create your own assays.  View the full product listing and conjugation protocol.
  • Compatible with standard flow cytometers
  • Free data analysis software
  • Excellent assay performance
  • Outstanding pricing

Currently available reactivitiesHuman, Mouse, Rat, Non-Human Primates
Maximum number of analytes in a plex13
Minimum required sample volume12.5 µl serum/plasma per well, 25 µl for tissue culture samples
Instrument requiredFlow cytometer capable of detecting PE and APC (or PerCP/FL3 on instruments without a red laser)
Tests per kit100 tests
Example Data

Now is your opportunity to get hands on training on how to perform the assays and analyze your data using LEGENDplex™ products with the scientists who developed the assays!


  • One or two days of training, depending on your schedule, at BioLegend headquarters in sunny San Diego. 
  • Hands on product usage at the bench, including setup and sample acquisition on flow cytometers, as well as data analysis using LEGENDplex™ software.
  • Plenty of time for troubleshooting and discussions with product development scientists.
  • Bringing your own samples is highly encouraged.
  • There is no charge for the training.
  • Housing and meals are not provided. Customers are responsible for their own travel and housing accommodations.
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