Intracellular Staining (ICS) with FACS analysis
Intracellular cytokine staining (ICS) combined with fluorescence‑activated cell sorting (FACS) analysis is a multiparametric flow‑cytometry‑based technique used to characterize and quantify antigen‑specific cellular immune responses at the single‑cell level. The assay enables the detection of intracellular cytokines and effector molecules following antigen‑specific stimulation. Cells are fixed and permeabilized to allow fluorochrome‑conjugated antibodies to bind intracellular targets and are subsequently analyzed by flow cytometry, where laser excitation and fluorescence detection permit identification and quantification of responsive cell populations.
ICS‑FACS provides high analytical sensitivity and allows simultaneous assessment of multiple functional markers together with surface phenotypic markers, enabling detailed characterization of immune cell subsets and polyfunctional responses. Commonly measured intracellular markers in VisMederi include interferon‑gamma (IFN‑γ), interleukin‑2 (IL‑2), interleukin‑4 (IL‑4), interleukin‑5 (IL‑5), tumor necrosis factor‑alpha (TNF‑α), and granzyme B. While the method offers comprehensive functional and phenotypic profiling, it requires careful assay optimization, appropriate controls, and specialized expertise for sample preparation and data analysis. VisMederi specializes in ICS and FACS-based immune monitoring and assay standardization (Harmonization and qualification of intracellular cytokine staining to measure influenza-specific CD4+ T cell immunity within the FLUCOP consortium; Harmonization and qualification of intracellular cytokine staining to measure influenza-specific CD4+ T cell immunity within the FLUCOP consortium.; Flow cytometry as an integrative method for the evaluation of vaccine immunogenicity: A validation approach – PMC), with extensive expertise in the characterization of antigen-specific cellular immune responses to respiratory viruses and vector-borne pathogens. Leveraging robust assay development and validation capabilities, VisMederi can adapt and implement customized cellular immunology assays for a wide range of viral and bacterial targets to support vaccine and therapeutic development programs.
