T helper (Th) cells play important roles in regulating immune responses. They secrete cytokines to stimulate various effector cells, such as cytotoxic T cells, B cells and macrophages. These cytokines are also involved in inflammatory and autoimmune diseases. Accurate measurement of Th cytokine expression is critical for in-depth understanding of the immune responses and many disease processes.
The LEGENDplex™ Human Th Cytokine Panel is a bead-based multiplex assay panel, using fluorescence–encoded beads suitable for use on various flow cytometers. This panel allows simultaneous quantification of 3 human cytokines, including IL-9, IL-10, TNF-α, which are collectively secreted by Th9. This assay panel provides higher sensitivities and broader dynamic range than traditional ELISA methods. The panel has been validated for use on serum and cell culture supernatant samples.
- Kit Contents
- Setup Beads 1: FITC Beads (1 mL)
- Setup Beads 2: PE Beads (1 mL)
- Setup Beads 3: Raw Beads (2 mL)
- Capture Beads (varies, see manual)
- Detection Antibodies (3.5 mL)
- Standard Cocktail, Lyophilized (1 vial)
- SA-PE (3.5 mL)
- Matrix B, Lyophilized (1 vial)
- Assay Buffer (25 mL)
- Wash Buffer, 20X (25 mL)
- Plate Sealers (4 sheets)
- Data Analysis Software Dongle
- V-bottom Plate
- Human Th Panel Manual
Learn more about LEGENDplex™ at biolegend.com/legendplex
Download the LEGENDplex™ software here.
- Materials Not Included
- Flow Cytometer
- Pipettes and Tips
- Reagent Reservoirs for Multichannel Pipettes
- Polypropylene Microfuge Tubes
- Vortex Mixer
- Aluminum Foil
- Absorbent Pads or Paper Towels
- Plate Shaker
- Tabletop Centrifuges
- A Vacuum Filtration Unit and a Vacuum Source (if using filter plates)
- 1.1 mL polypropylene micro FACS tubes, in 96-tube rack
- If I don't have a vacuum, how do I remove the liquid from my plate?
If you do not have a vacuum, the assay should be run in a V-bottom plate. After centrifugation using a swinging-bucket rotor with a plate adaptor, you can remove the liquid by flicking the plate quickly, dumping the contents into a sink, and patting it dry carefully on a stack of clean paper towels without losing the beads. Alternatively, you can remove the liquid by using a pipette.
- Should I perform the assay with the filter plates or with V-bottom plates?
Filter plates or V-bottom plates have been included in some kits for your convenience. A vacuum filtration unit is required to work with the filter plates. However, if you don’t have access to a vacuum manifold or if you prefer, then you can use the V-bottom plates and follow the recommended assay protocols for the type of plates you choose. All plates should be made from low binding polypropylene. Polystyrene ELISA or cell culture plates should not be used.
- After I finish the staining process, how long can I wait before reading my LEGENDplex™ samples?
The samples can be kept overnight at 4°C while being protected from exposure to light and be read the next day. There may be a decrease in signal, but overall, the assay results should not be affected. Storing the samples for extended periods of time is not recommended, as it could lead to further reductions in signal.
- What is the shelf life of LEGENDplex™ kits?
LEGENDplex™ kits are guaranteed for 6 months from the date of receipt, but may have a shelf life of up to 2 years from the date of manufacture.
- Is special software required for data analysis?
Typically flow cytometers generate output files in FCS format (e.g. FCS 2.0, 3.0, or 3.1) and in some cases in list mode file format (LMD). Other software may be available to analyze FCS files. Data generated using LEGENDplex™ kits can be analyzed using the freely available LEGENDplex™ data analysis software. Please check our website for the most updated versions of the software.