Journal:

Article Title: Microsatellite Analysis of Hereditary Nonpolyposis Colorectal Cancer-Associated Colorectal Adenomas by Laser-Assisted Microdissection

doi:

Figure Lengend Snippet: Patient Characteristics

Article Snippet: The slides were incubated overnight at 4°C with primary antibody against MSH2 (clone FE11, dilution 1:100; Oncogene Sciences, Cambridge, MA), MLH1 (clone G168-728, dilution 1:200; PharMingen, San Diego, CA), or MSH6 (clone 44, dilution 1:600; Becton Dickinson Transduction Laboratories, San Diego, CA) proteins.

Techniques:

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