fix/perm buffer ebioscience foxp3/tf staining buffer set  (Thermo Fisher)

Journal: eBioMedicine

Article Title: Pharmacological inhibition of tyrosine protein-kinase 2 reduces islet inflammation and delays type 1 diabetes onset in mice

doi: 10.1016/j.ebiom.2025.105734

Figure Lengend Snippet: TYK2 inhibition reshapes innate and adaptive immunity in blood, spleen, and PLN of RIP-LCMV-GP mice . Peripheral blood mononuclear cells (PBMCs), splenocytes, and immune cells from PLN were isolated and labelled with CD11b + MHCII + CD11c + (dendritic cells, DCs), CD11b + MHCII + F4/80 + (macrophages), CD11b + CD49 + (mature NK cells), and CD11b + CD49 - (immature NK cells). Immune cell characterisation was performed using flow cytometry. (a – c) Percentage of DCs, macrophages, mature NK cells, and immature NK cells from 3, 7, and 14-days post-inoculation from vehicle (grey) and TYK2i-treated (red) RIP-LCMV-GP mice. ( d–f ) Percentage of CD4 + FoxP3 + CD25 + Tregs, CD4 + PD1 + T cells, and CD8 + PD1 + T cells from 3, 7, and 14-days post-inoculation from vehicle (grey) and TYK2i-treated (red) RIP-LCMV-GP mice. Data are presented as mean with 95% CI, and individual data points are included, n = 3–5 mice per condition; Statistical significance was determined by a Mann–Whitney U test.

Article Snippet: Single cell suspensions were permeabilised for intracellular staining using FoxP3 fix/perm buffer (Thermo Fisher Scientific, catalogue #00-5523-00) at 4 °C for 20 min, followed by a wash with permeabilisation buffer.

Techniques: Inhibition, Isolation, Flow Cytometry, MANN-WHITNEY

Journal: eBioMedicine

Article Title: A probiotic approach identifies a Treg-centred immunoregulation via modulation of gut microbiota metabolites in people with multiple sclerosis and healthy individuals

doi: 10.1016/j.ebiom.2025.105743

Figure Lengend Snippet: Probiotic supplementation shifts the peripheral T cell composition towards an anti-inflammatory phenotype in HC and pwMS. PBMCs from HC and pwMS were analysed for their respective marker expression via flow cytometry before (baseline), after two and six weeks of probiotic supplementation and six weeks after termination of probiotic intake (WO). The graphs show relative changes of the analysed cell population normalised to each individual's baseline. Black lines represent individual study participants. Orange lines indicate the median change. (A, B) Two weeks of probiotic intake did not significantly change IL-17A+ Th17 cells in HC (A) and pwMS (B). (C, D) IFNγ+ Th1 cells revealed no effect in HC (C) but a decrease in pwMS (D) after two weeks. (E–G) FoxP3+ Treg cells increased in HC (E) and pwMS (F). (G) Representative dot plots of FoxP3+ Treg cells demonstrate the increase after two weeks of probiotic intake (right) compared to baseline (left). (H, I) Six weeks of probiotic intake did not change significantly Th1 cell frequencies in HC (H) but decreased Th1 cells in pwMS (I). (J, K) Treg cell frequencies were significantly increased in HC (J) and pwMS (K) after six weeks. (L–O) Th1 and Treg cell frequencies were analysed 6 weeks after the last probiotic intake (washout, WO). Relative frequencies of Th1 cells reduced in HC (L) and pwMS (M), whereas Treg cell frequencies remained unchanged in HC (N) but dropped in pwMS (O) compared to baseline frequencies. Significance for changes over time were calculated using unnormalised cell frequencies with Wilcoxon matched-pairs signed rank test. (A) n = 41, p = 0.672. (B) n = 28, p = 0.333. (C) n = 40, p = 0.952. (D) n = 28, ∗p = 0.029. (E) n = 37, ∗∗p = 0.005. (F) n = 28, ∗p = 0.036. (H) n = 17, p = 0.963. (I) n = 15, ∗p = 0.0302. (J) n = 17, ∗p = 0.0295. (K) n = 15, ∗p = 0.0129. (L) n = 17, p = 0.404. (M) n = 15, p = 0.064. (N) n = 17, p = 0.382. (O) n = 15, p = 0.629.

Article Snippet: Cells were then fixed and made permeable using the FOXP3 Fix/Perm Buffer Set (eBioscience) according to the manufacturer's instructions.

Techniques: Marker, Expressing, Flow Cytometry