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FluoroSpot MAX™ Deluxe Set Human IFN-γ/IL-6

Pricing & Availability
Regulatory Status
RUO
Other Names
IFNG, IFG, IFI, Interferon gamma, Immune interferon, Interleukin 6, B-cell Stimulatory Fa
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publications
FluoroSpot_Human-IFN-Y_IL-6_073025
Human PBMC (5,000 cells/well) were incubated for 5 hours in the absence or presence of either 1X Cell Activation Cocktail without Brefeldin A (BioLegend, Catalog No. 423301) or 1X Cell Activation Cocktail without Brefeldin A (Cat. No. 423301) or 1X Cell Activation Cocktail with Brefeldin A (Cat. No. 423303) or in the presence of 100 ng/mL LPS. All figures showed the merged figure of IFN-γ (PE, Yellow) and IL-6 (APC, Red). BioLegend’s Cell Activation cocktail contains PMA and ionomycin at the optimized concentration. Brefeldin A functions to inhibit intracellular protein transport, as a result, it inhibits secretion of cytokines such as IFN-γ and IL-6. The number of cells secreting Human IFN-γ/IL-6 was analyzed by FluoroSpot and quantified using an image cytometer equipped with the required fluorescence filters.
  • FluoroSpot_Human-IFN-Y_IL-6_073025
    Human PBMC (5,000 cells/well) were incubated for 5 hours in the absence or presence of either 1X Cell Activation Cocktail without Brefeldin A (BioLegend, Catalog No. 423301) or 1X Cell Activation Cocktail without Brefeldin A (Cat. No. 423301) or 1X Cell Activation Cocktail with Brefeldin A (Cat. No. 423303) or in the presence of 100 ng/mL LPS. All figures showed the merged figure of IFN-γ (PE, Yellow) and IL-6 (APC, Red). BioLegend’s Cell Activation cocktail contains PMA and ionomycin at the optimized concentration. Brefeldin A functions to inhibit intracellular protein transport, as a result, it inhibits secretion of cytokines such as IFN-γ and IL-6. The number of cells secreting Human IFN-γ/IL-6 was analyzed by FluoroSpot and quantified using an image cytometer equipped with the required fluorescence filters.
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Description

Human IL-6 (Interleukin), also known as B-cell Stimulatory Factor 2 (BSF2), Interferon beta-2 (IFN-β-2), Hybridoma Growth Factor (HGF), is a cytokine, which is expressed by many different cell types such as macrophages, T-cells, osteoblasts, endothelial cells, and hepatocytes. It is involved in both pro-inflammatory and anti-inflammatory responses. In particular, IL-6 has critical roles in multiple different biological processes including but not limited to B-cell maturation, osteoclast formation, nerve cell differentiation, muscle healing and hypertrophy. Thus, abnormal level of expression and activity of IL-6 can result in a wide range of diseases from chronic inflammation, cancer, to neurodegenerative diseases. Human IFN-γ (Interferon-gamma) is a multifunctional cytokine which is the sole member of the class II Interferon. Although a variety of immune cells can express IFN-γ, most of IFN-γ is produced by T cells and NK cells. It has critical roles in many processes including, but not limited to, anti-viral response, regulating Th1/Th2 balance, enhancing antigen presentation, and controlling cellular proliferation and apoptosis. IFN-γ can bind to 2 receptors: IFNGR1 and IFNGR2, which transduce the signal downstream through the JAK-STAT pathway. Because of its multifunctional nature, dysregulation of IFN-γ expression and activity has been implicated in many diseases such as autoimmune and cancer.

BioLegend’s FluoroSpot MAX™ Deluxe Set contains pre-optimized essential reagents and buffers for the simultaneous detection of human IL-6 and IFN-γ secreting cells. In this assay, anti-human IL-6 and IFN-γ capture antibodies are coated on a clear-bottom microplate. Cells (stimulated or unstimulated) are added to the wells and incubated, allowing secreted IL-6 and/or IFN-γ to bind to nearby capture antibodies. After incubation, cells are removed, and fluorescent conjugated detection antibodies are added. The resulting spots represent the "footprints" of individual IL-6 and IFN-γ-secreting cells, enabling precise quantification of both single cytokine-producing cells and dual cytokine-producing cells within the sample.

Product Details
Technical Data Sheet (pdf)

Kit Details

Kit Contents
  • Human IL-6 (2-plex) Capture Antibody (200X) 
  • Human IFN-γ (2-plex) Capture Antibody (200X) 
  • APC Human IL-6 Detection Antibody (200X)  
  • PE Human IFN-γ Detection Antibody (200X)  
  • Human IFN-γ Standard (used as a positive control) 
  • Human IL-6 Standard (used as a positive control) 
  • Coating Buffer A (5X) 
  • Assay Diluent B (5X)
Materials Not Included
  • Flat Bottom High Bind Microplate: Corning® Clear Bottom Black Polystyrene Microplates, 96 Well (Catalog No. 3601) is recommended, or equivalent.
  • Black Microplate Lid: Corning® Universal Black Microplate Lid with Corner Notch (Catalog No. 3935) is recommended, or equivalent
  • Wash Buffer: BioLegend Cat. No. 421601 is recommended, or PBS + 0.05% Tween-20  
  • Plate Sealers: BioLegend Cat. No. 423601 is recommended 
  • 0.22 μm filters
  • Sterile PBS (Phosphate-Buffered Saline): 8.0 g NaCl, 1.16 g Na2HPO4, 0.2 g KH2PO4, 0.2 g KCl, add deionized water to 1 L; pH to 7.4, 0.22 μm filtered
  • Sterile cell culture media
  • 37°C CO2 incubator
  • Deionized (DI) water 
  • Adjustable pipettes to measure volumes ranging from 2 μL to 1 mL 
  • Wash bottle, manifold dispenser, or automated microplate washer 
  • Tubes for dilution
  • Timer
  • Alcohol Wipes 
  • Image cytometer or other imaging platform capable of whole-well imaging. The device should be equipped with the appropriate filters for the fluorophores utilized in each assay.
  • Optional: Carboxyl Beads B4 (Cat. No. 740175 or equivalent)

Product Details

Verified Reactivity
Human
Application

FluoroSpot

Antigen Details

Cell Type
Endothelial cells, Macrophages, NK cells, NKT cells, Osteoblasts, T cells, Th1
Biology Area
Cancer Biomarkers, Cell Biology, Immuno-Oncology, Immunology, Innate Immunity, Neurodegeneration, Signal Transduction
Molecular Family
Cytokines/Chemokines, Innate Immune Signaling
Gene ID
3458; 3569 View all products for this Gene ID
UniProt
View information about IFN-gamma/IL-6 on UniProt.org

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Go To Top Version: 1    Revision Date: 07/17/2025

For Research Use Only. Not for diagnostic or therapeutic use.

 

This product is supplied subject to the terms and conditions, including the limited license, located at www.biolegend.com/terms) ("Terms") and may be used only as provided in the Terms. Without limiting the foregoing, BioLegend products may not be used for any Commercial Purpose as defined in the Terms, resold in any form, used in manufacturing, or reverse engineered, sequenced, or otherwise studied or used to learn its design or composition without express written approval of BioLegend. Regardless of the information given in this document, user is solely responsible for determining any license requirements necessary for user’s intended use and assumes all risk and liability arising from use of the product. BioLegend is not responsible for patent infringement or any other risks or liabilities whatsoever resulting from the use of its products.

 

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