eipa  (MedChemExpress)

Journal: bioRxiv

Article Title: Distinct sensing of BCAAs by mTOR and c-Myc governs T cell proliferation, independent of catabolism

doi: 10.64898/2026.01.16.699967

Figure Lengend Snippet: (A) Schematic of Day 0-3 BCAA restriction and addback culture. Bulk CD8 T cells were isolated and purified from spleens of WT mice, activated with anti-CD3 and anti-CD28 antibodies (2ug/mL each) for 3 days in BCAA-free TCM with addback of varying combinations of Leu, Ile and Val (0.8mM or 2.4mM [3X] each as indicated). (B) Frequency of viable CD8 T cells after Day 0-3 BCAA culture (2.4mM addbacks). (C) Frequency of dividing cells after Day 0-3 BCAA culture (% of cells with CTV dilution, CTV stain performed on Day 0). (D) Frequency and representative flow plots of cytokine-producing CD8 T cells after Day 0-3 BCAA culture (2.4mM addbacks). (E) Schematic of Day 2-3 BCAA restriction with EIPA and addback culture. Bulk CD8 T cells were isolated and purified from spleens of WT mice, activated with anti-CD3 and anti-CD28 antibodies (2ug/mL each) for 2 days in replete TCM followed by 1 day in BCAA-free TCM with addback of varying combinations of Leu, Ile and Val (0.8mM each) and simultaneous treatment with 25uM EIPA or vehicle control. (F) Frequency of highly dividing (>4 divisions) cells as measured by CTV dilution (CTV stain performed immediately after CD8 purification on Day 0 of culture). All error bars are representative of 3 technical replicates. Statistical significance in (F) was calculated using two-way ANOVA with multiple comparisons and Sidak’s correction. ****p<0.0001.

Article Snippet: For all experiments using pharmacological agents, the following doses were used unless otherwise noted: EIPA (25uM, MedChem Express), Cycloheximide (50ug/mL, Sigma), MG132 (10uM, Sigma), Torin (1uM, Cayman), ISRIB (1uM, Ambeed), Rapamycin (50nM, Sigma).

Techniques: Isolation, Purification, Staining, Control

Journal: Nature Communications

Article Title: Chimeras co-targeting antigens and FcγRIIb trigger degradation of extracellular soluble proteins and pathological aggregates

doi: 10.1038/s41467-025-67207-4

Figure Lengend Snippet: a , b Effect of inhibitors targeting different endocytic pathways on FcRTAC internalization. Huvec-2b cells were pre-treated with endocytosis inhibitors for 1 h, followed by the addition of pH-sensitive reagent-prelabeled NK-2-12-SK3 ( a ) or NK-2-12-SK4 ( b ). Fluorescence changes were monitored hourly over 12 h. Chlorpromazine: clathrin-mediated endocytosis inhibitor; cytochalasin D: macropinocytosis and phagocytosis inhibitor; amiloride (EIPA): macropinocytosis inhibitor; nystatin: caveolae pathway inhibitor. c – e , Live-cell imaging of FcRTAC-mediated IgE endocytosis. Cy3-labeled IgE (red) was incubated with Huvec-2b cells in the presence or absence of Cy5-labeled FcRTACs (white) for 2 h. Cells were stained with LysoTracker (green) and Hoechst (blue) for confocal imaging. Images were acquired using a 40× objective on a confocal microscope ( c , scale bar: 20 μm) or a super-resolution microscope ( d and e ; scale bar: 10 μm). f Subcellular distribution of FcγRIIb and FcRTACs in Huvec-2b cells. Huvec-2b cells were incubated with Cy5-labeled FcRTACs (red) for 1 h. After fixation and permeabilization, cells were stained for Lamp-1 (green) and FcγRIIb (white) using primary antibodies and their corresponding fluorescent secondary antibodies. Nuclei were stained by DAPI (blue). Images were acquired using a 40× objective on a confocal microscope. Scale bar: 10 μm. c–f Experiments were performed in triplicate and repeated three times with similar results.

Article Snippet: The inhibitors included Chlorpromazine (Sigma, 31679), Cytochalasin D (MCE, HY-N6682), Amiloride (MCE, HY-101840), and Nystatin (Solarbio, N8040).

Techniques: Fluorescence, Live Cell Imaging, Labeling, Incubation, Staining, Imaging, Microscopy, Super-Resolution Microscopy