ArticleClinical and diagnostic laboratory immunology2002
Differential modulation of surface and intracellular protein expression by T cells after stimulation in the presence of monensin or brefeldin A.
Article in Clinical and diagnostic laboratory immunology, 2002. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 39 papers.
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Who cites it
39 citing papers in PubMed, 69 citations in OpenAlex.
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- Maximizing Insights, Minimizing Animal Testing: A Framework for Validating Multiparametric Single-Cell Cytokine Analysis Panels.European journal of immunology · 2025Article
- Effective and Successful Quantification of Leukemia-Specific Immune Cells in AML Patients' Blood or Culture, Focusing on Intracellular Cytokine and Degranulation Assays.International journal of molecular sciences · 2024Article
- High-dimensional multi-pass flow cytometry via spectrally encoded cellular barcoding.Nature biomedical engineering · 2024Article
- Spatially Defined Cell-Secreted Protein Detection Using Granular Hydrogels: μGeLISA.ACS biomaterials science & engineering · 2023Article
- An optimized flow cytometry protocol for simultaneous detection of T cell activation induced markers and intracellular cytokines: Application to SARS-CoV-2 immune individuals.Journal of immunological methods · 2023Article
- A whole blood intracellular cytokine assay optimised for field site studies demonstrates polyfunctionality of CD4+ T cells in acute scrub typhus.PLoS neglected tropical diseases · 2023Article
- A novel phosphocholine-mimetic inhibits a pro-inflammatory conformational change in C-reactive protein.EMBO molecular medicine · 2023Article
- Influenza-Induced CD103ImmunoHorizons · 2022Article
- IL-36γ is secreted through an unconventional pathway using the Gasdermin D and P2X7R membrane pores.Frontiers in immunology · 2022Article
- Lymphostatin, a virulence factor of attaching and effacingFrontiers in cellular and infection microbiology · 2022Article
- Guidelines for the use of flow cytometry and cell sorting in immunological studies (third edition).European journal of immunology · 2021Review
- Measuring single-cell protein secretion in immunology: Technologies, advances, and applications.European journal of immunology · 2021Review
- Article
- Optimized Intracellular Staining Reveals Heterogeneous Cytokine Production Ability of Murine and Human Hematopoietic Stem and Progenitor Cells.Frontiers in immunology · 2021Article
- Exosomal delivery of NF-κB inhibitor delays LPS-induced preterm birth and modulates fetal immune cell profile in mouse models.Science advances · 2021Article
Corrections and comments
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Authors and funding
2 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Intracellular cytokine staining is an increasingly popular analytical tool that can be used to define the profile of cytokines in various disease states. One important requirement for this assay is the inclusion of a protein transport inhibitor in stimulated cell cultures to trap the cytokine, thus allowing a brighter signal. Two compounds commonly used for this purpose are brefeldin A (BFA) and monensin (MN). Flow cytometry was used to assess the differential effects of BFA and MN on surface CD3, -4, -8, and -69 expression and the intracellular expression of gamma interferon (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) following stimulation with phorbol myristate acetate and ionomycin. We found that BFA blocked the majority of CD3(+) cells from expressing surface CD69, but BFA did not inhibit intracellular CD69 expression. MN did not significantly inhibit surface CD69 expression. With regard to lymphocyte marker expression following activation, surface CD4 expression was significantly downregulated; however, less downregulation was observed with BFA treatment than with MN treatment. Analyzing intracellular cytokine expression, BFA trapped a greater percentage of TNF-alpha inside activated cells than MN. An analysis of the cytokine concentration in culture supernatants indicated that cells treated with MN released TNF-alpha and IFN-gamma from the cells, while the BFA-treated cells released IFN-gamma only. With prolonged (18-h) stimulation, the cells treated with MN were less viable than those treated with BFA. We conclude that the choice of a protein transport inhibitor is an important variable in this assay. When developing this method as a tool for clinical immunology laboratory analysis, investigators should consider the differential effects of BFA and MN on results.
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