Journal: Oncotarget

Article Title: New strategy to rescue the inhibition of osteogenesis of human bone marrow-derived mesenchymal stem cells under oxidative stress: combination of vitamin C and graphene foams

doi: 10.18632/oncotarget.12456

Figure Lengend Snippet: ( A ) Representative image of BM-MSCs 5 days after seeding. ( B – D ) Characterization of BM-MSCs by flow cytometry. The majority of the cells are CD105 + , CD44 + and CD29 + , which are typical characteristic phenotypes of BM-MSCs. ( E ) BM-MSC can be differentiated into adipogenic and myogenic lineages. Adipogenic differentiation was characterized by Oil Red O staining, while myogenic differentiation was evidenced by the formation of myotubes stained with crystal violet. ( F ) Representative image of immunostaining of BM-MSCs on GF scaffold, stained by anti-β-tubulin (green) and DAPI for nucleus (blue). Effects of different concentrations of VC (5 to 100 μg/ml, 5 days) ( G ) and H 2 O 2 exposure (0.1 to 2 mM, 24 hours) ( H ) on cell viability of BM-MSCs cultured for 5 days, measured by MTT assay. ( I ) Cell viability of BM-MSCs in the five experimental groups. Data were presented as mean ± SEM. * p < 0.05 vs control.

Article Snippet: Antibodies used in the study were as follows: CD105-FITC (MCA1557FT, Serotec, Oxford, UK), CD44-FITC (MCA643FA, Serotec), and CD29-FITC (MCA1949FT, Serotec).

Techniques: Flow Cytometry, Staining, Immunostaining, Cell Culture, MTT Assay, Control

Journal: bioRxiv

Article Title: CLIC-dependent internalization of caveolin-1 to lysosomal vacuoles in response to osmotic regulation

doi: 10.1101/2025.08.26.672461

Figure Lengend Snippet: A) MDA-MB-231 cells were incubated in isotonic media, in hypotonic media for 5, 15, 30 and 60 minutes or under Recovery conditions in which hypotonic media was replaced after 60 minutes with isotonic media for 5, 15, 30 or 60 minutes. Cells were labeled for Cav1, fixed and imaged by TIRF widefield imaging. Average Cav1 intensity per cell was quantified and normalized to isotonic control. Representative images of select conditions are shown. (n=3 independent experiments with >30 cells for each condition; ANOVA with Dunnett’s post-test comparing to isotonic control; **p < 0.01; ***p < 0.001; Scale bar: 2µm). B) STED imaging of anti-Cav1 labeling MDA-MB-231 cells incubated in isotonic media or in hypotonic media for 15 or 60 minutes. Cav1 endocytic vacuoles were counted per cell. (n>28 from three independent experiments for isotonic and hypotonic 15 minutes, four independent experiments for other conditions; (ANOVA with Dunnett post-test comparing to isotonic control; *p < 0.05; ****p < 0.0001; Scale bar: 4µm; Inset scale bar: 1µm). C) MDA-MB-231 cells incubated with isotonic or hypotonic media for 60 minutes were fixed and labeled for Cav1, CD44 and β1-integrin (Scale bar: 4µm). The diameter of individual Cav1, CD44, and β1-integrin positive endosomes was measured from three independent experiments.

Article Snippet: Primary antibodies: rabbit anti-Cav1 mAb (3267) was purchased from Cell Signaling Technology, rat anti-CD44 mAb (22530) from Novus, and mouse anti-β1 integrin mAb (sc-53711) from Santa Cruz.

Techniques: Incubation, Labeling, Imaging, Control

Journal: bioRxiv

Article Title: CLIC-dependent internalization of caveolin-1 to lysosomal vacuoles in response to osmotic regulation

doi: 10.1101/2025.08.26.672461

Figure Lengend Snippet: A) MDA-MB-231 cells were incubated with anti-CD44 and anti-β1 integrin antibodies in isotonic media for 60 minutes at 4°C or 37°C or in hypotonic media for 5, 15, 30 or 60 minutes at 37°C. As indicated, an acid wash treatment was performed at 4°C to remove surface antibodies and cells were fixed and labeled with secondary antibodies to anti-CD44 and anti-β1 integrin as well as for endogenous Cav1. (n= >30 cells per condition from three independent experiments; ANOVA with Dunnett post-test comparing each condition to isotonic 4°C control; *p < 0.05; ***p < 0.001; ****p < 0.0001; Scale bars: 2 µm). B) MDA-MB-231 cells were transfected with wild-type CDC42-GFP or dominant negative (Dom Neg) CDC42-GFP, incubated with isotonic or hypotonic media for 60 minutes and then fixed and labeled for Cav1 and CD44. CDC42-GFP was also imaged and the number of Cav1 endocytic vacuoles counted per cell (n>30 cells from three independent experiments; ANOVA with Dunnett’s post-test comparing to isotonic control; ****p < 0.0001; Scale bar: 2µm).

Article Snippet: Primary antibodies: rabbit anti-Cav1 mAb (3267) was purchased from Cell Signaling Technology, rat anti-CD44 mAb (22530) from Novus, and mouse anti-β1 integrin mAb (sc-53711) from Santa Cruz.

Techniques: Incubation, Labeling, Control, Transfection, Dominant Negative Mutation

Journal: bioRxiv

Article Title: CLIC-dependent internalization of caveolin-1 to lysosomal vacuoles in response to osmotic regulation

doi: 10.1101/2025.08.26.672461

Figure Lengend Snippet: A) MDA-MB-231 cells were transfected with LAMP-GFP, incubated with isotonic or hypotonic media for 60 minutes and then fixed and labeled for Cav1. LAMP-GFP was also imaged and the number of Cav1 endocytic vacuoles counted per cell (n>30 from cells three independent experiments with >30 cells for each condition; ANOVA with Dunnett’s post-test comparing to isotonic control; ****p < 0.0001; Scale bar: 2 µm). B) Cav1 CRISPR knockout MDA-MB-231 cells were transfected with Cav1-WT-HA or Cav1-K*R-HA lysine mutant, incubated with isotonic or hypotonic media for 15 or 60 minutes and then fixed and labeled for Cav1 and CD44. (n>29 cells from three independent experiments; ANOVA with Tukey post-test comparing WT to K*R at the same isotonic or hypotonic conditions and the respective hypotonic condition to isotonic control; Scale bar: 2µm). C) MDA-MB-231 cells were transfected with LAMP1-mScarlet and Cav1-Halo, where indicated pre-treated with 100nM BafA for 2 hours, labeled with Halo-LIVE RED for 30 minutes and then incubated for 30 minutes with 1uM LysoSensor green and LIVE RED Halotag ligand. Cells were then placed in isotonic or hypotonic imaging media and multiple cells imaged over a period of 30 minutes in isotonic DMEM or after 30 minutes of hypotonic shock. LysoSensor and Cav1 intensity within LAMP1-positive lysosomes were quantified. (n>38 cells from three independent experiments; ANOVA with Tukey’s post-test; *p < 0.05; ****p < 0.0001; Scale bar: 2 µm).

Article Snippet: Primary antibodies: rabbit anti-Cav1 mAb (3267) was purchased from Cell Signaling Technology, rat anti-CD44 mAb (22530) from Novus, and mouse anti-β1 integrin mAb (sc-53711) from Santa Cruz.

Techniques: Transfection, Incubation, Labeling, Control, CRISPR, Knock-Out, Mutagenesis, Imaging

Journal: bioRxiv

Article Title: CLIC-dependent internalization of caveolin-1 to lysosomal vacuoles in response to osmotic regulation

doi: 10.1101/2025.08.26.672461

Figure Lengend Snippet: A) Cell volume of MDA-MB-231 and PC3 cells switched from isotonic to hypotonic media was measured by live phase imaging. Black line indicates addition of hypotonic media (n=3 independent experiments; each experiment includes 2-3 technical replicates and each technical replicate consists of 9 ROIs). B) MDA-MB-231 and PC3 cells were incubated in isotonic media, or hypotonic or hypertonic media for 5, 15, 30 and 60 minutes. Cells were labeled for Cav1 and CD44 and Cav1 endocytic vacuoles counted per cell. Representative images for select time points are shown. (n>28 cells from three independent experiments; ANOVA with Dunnett’s post-test comparing each condition to isotonic control; *p < 0.05; **p < 0.01; ****p < 0.0001; Scale bar: 2 µm). C) PC3 cells and PC3 cells stably transfected Cavin-1-GFP were incubated in isotonic, hypotonic and hypertonic shock media for 60 minutes and then fixed and labeled for Cav1. Cavin-1-GFP was imaged in the GFP channel and Cav1 endocytic vacuoles counted per cell. (n>28 cells from three independent experiments; ANOVA with Tukey’s post-test and comparing PC3 to PC3+Cavin-1 and each condition with the respective isotonic control; **p < 0.01; ***p < 0.001; ****p < 0.0001; Scale bar: 2 µm).

Article Snippet: Primary antibodies: rabbit anti-Cav1 mAb (3267) was purchased from Cell Signaling Technology, rat anti-CD44 mAb (22530) from Novus, and mouse anti-β1 integrin mAb (sc-53711) from Santa Cruz.

Techniques: Imaging, Incubation, Labeling, Control, Stable Transfection, Transfection

Journal: eLife

Article Title: Inflammasome activation leads to cDC1-independent cross-priming of CD8 T cells by epithelial cell-derived antigen

doi: 10.7554/eLife.72082

Figure Lengend Snippet: ( A ) Overview of experimental setup for analyzing OT-I responses to OvaFla production in IECs. ( B ) Quantification of OT-Is as a percent of total CD8 + T cells per spleen (left) and mesenteric lymph node (right). ( C ) Total number of OT-Is per spleen (left) and mesenteric lymph node (right). ( D ) Total number of CD62L – CD44 + OT-Is per spleen (left) and mesenteric lymph node (right). Samples with fewer than 20 OT-Is were excluded from CD62L, CD44 calculations. Tissues were harvested and analyzed at day 5 post tamoxifen chow start. Data are pooled from three biological replicates, and each dot represents an individual mouse. Data shown as mean ± SD. Significance calculated using one-way ANOVA and Tukey’s multiple comparisons test (*p < 0.05, **p < 0.01, ***p < 0.001). Only p values between wild-type (WT) and other experimental groups are shown. See for exact p values. Figure 3—source data 1. Statistical data for .

Article Snippet: Antibody , Anti-mouse CD44 BB515 (rat monoclonal) , BD , Clone: IM9; Cat#: 564587 , FC(1:300).

Techniques:

Journal: eLife

Article Title: Inflammasome activation leads to cDC1-independent cross-priming of CD8 T cells by epithelial cell-derived antigen

doi: 10.7554/eLife.72082

Figure Lengend Snippet: ( A ) Schematic depicting the production and analysis workflow of chimeric bm1 + OvaFla mice (left). At the right, an illustration of either wild-type (WT) OvaFla mice (left of the dashed line) or Nlrc4 –/– OvaFla mice (right of the dashed line) following lethal irradiation and reconstitution with bone marrow from B6.SJL mice. ( B ) Quantification of OT-Is as a percent of total CD8 + T cells (left), the total number of OT-Is (middle), and the total number of CD62L – CD44 + OT-Is (right) in the spleen. ( C ) Quantification of OT-Is as a percent of total CD8 + T cells (left), the total number of OT-Is (middle), and the total number of CD62L – CD44 + OT-Is (right) in the mesenteric lymph nodes. Tissues were harvested and analyzed at day 5 post tamoxifen chow start. (B–C) Data are pooled from three biological replicates, and each dot represents an individual mouse. Data shown as mean ± SD. Significance calculated using one-way ANOVA and Tukey’s multiple comparisons test (*p < 0.05, **p < 0.01, ***p < 0.001). Only p values between WT and other experimental groups are shown. See for exact p values. Figure 4—source data 1. Statistical data for .

Article Snippet: Antibody , Anti-mouse CD44 BB515 (rat monoclonal) , BD , Clone: IM9; Cat#: 564587 , FC(1:300).

Techniques: Irradiation

Journal: eLife

Article Title: Inflammasome activation leads to cDC1-independent cross-priming of CD8 T cells by epithelial cell-derived antigen

doi: 10.7554/eLife.72082

Figure Lengend Snippet: ( A ) Percent of CD45 + cells that are conventional type one dendritic cells (cDC1s) in bm1 chimera mice that received either Batf3 + or Batf3 – donor bone marrow. ( B ) Quantification of OT-Is as a percent of total CD8 + T cells (top), the total number of OT-Is (middle), and the total number of CD62L – CD44 + OT-Is (bottom) in the spleen. ( C ) Quantification of OT-Is as a percent of total CD8 + T cells (top), the total number of OT-Is (middle), and the total number of CD62L – CD44 + OT-Is (bottom) in the mesenteric lymph nodes. ( D ) Quantification of OT-Is that have out-diluted the CellTrace Violet dye in the spleen (top) and mesenteric lymph nodes (bottom). ( E ) Percent of CD62L – CD44 + OT-Is in the mesenteric lymph node that are CCR9 + . Tissues were harvested and analyzed at day 5 post tamoxifen chow start. Samples with fewer than 20 OT-Is were excluded from CD62L, CD44, and CCR9 calculations. (A) Data are from a single experiment. (B–D) Data are pooled from three biological replicates. (E) Data are pooled from two biological replicates. Each dot represents an individual mouse. Data shown as mean ± SD. Significance calculated using one-way ANOVA and Šídák’s multiple comparisons test (*p < 0.05, **p < 0.01, ***p < 0.001). See for exact p values. Figure 5—source data 1. Statistical data for .

Article Snippet: Antibody , Anti-mouse CD44 BB515 (rat monoclonal) , BD , Clone: IM9; Cat#: 564587 , FC(1:300).

Techniques:

Journal: eLife

Article Title: Inflammasome activation leads to cDC1-independent cross-priming of CD8 T cells by epithelial cell-derived antigen

doi: 10.7554/eLife.72082

Figure Lengend Snippet:

Article Snippet: Antibody , Anti-mouse CD44 BB515 (rat monoclonal) , BD , Clone: IM9; Cat#: 564587 , FC(1:300).

Techniques: Generated, CRISPR, Purification, Enzyme-linked Immunosorbent Assay, Software

Journal: Molecular Cancer

Article Title: Molecular analysis reveals heterogeneity of mouse mammary tumors conditionally mutant for Brca1

doi: 10.1186/1476-4598-7-29

Figure Lengend Snippet: Characterization of cancer stem cell markers in Brca1 cell lines. Cell lines A1.1, B1.15, P2.1 and P3.17 were stained with fluorescently-conjugated mAb specific to CD44 (A) , or CD24 (B) , in parallel with isotype control-matched mAb and analyzed by flow cytometry. In the histograms shown, the thick black line represents positive staining and the gray filled lines represent the isotype control-matched mAb. (C) Bivariate plots represent the double staining of cell lines with fluorescently-conjugated CD44 and CD24 mAb. The quadrants are gated based on isotype controls and separate the double positive, single positive, and double negative populations. The percent of positive fraction for single label and CD44 + /CD24 - cells after compensation are indicated in each panel.

Article Snippet: Cells from each cell line were grown to 70% confluence, scraped or trypsinized, stained with PE Anti-mouse CD44 (BD Pharmingen, San Jose, CA), APC Anti-mouse CD24 (Biolegend, San Diego, CA), FITC Anti-mouse CD44 (Southern Biotech, Birmingham, AL), or APC Anti-mouse CD117 (Biolegend).

Techniques: Staining, Control, Flow Cytometry, Double Staining