rabbit anti cap1  (Proteintech)

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Shear stress induces monocyte/macrophage-mediated inflammation by upregulating cell-surface expression of heat shock proteins.

doi: 10.1016/j.biopha.2023.114566

Figure Lengend Snippet: Fig. 2. Effect of shear stress on surface proteins. (A) Serum-starved THP-1 cells were replaced with the culture medium and exposed to shear stress (SS) (12 dynes/ cm2) for 24 h. Surface proteins exposed to shear stress were eluted, resolved on a 10% gel with protein markers (M), and visualized after silver staining. The bands detected at high intensity in the 24 h SS lane in comparison with the 0-h SS lane were excised for mass spectrometry. The protein identification of the band is detailed in the Table.(B) Transcript levels of HSP105, HSP90, HSP70, and CAP1 were assessed using real-time PCR. **P < 0.01, vs. 0 h SS; *P < 0.05, vs. 0 h SS.(C and D) HSP105, HSP90, HSP70, and CAP1 proteins were measured by western blotting using (C) total cell lysates and (D) fraction of surface proteins.(E and F) After serum starvation, THP-1 cells were subjected to shear stress for 24 h. Cells were immunostained with HSP105, HSP90, HSP70, and CAP1 using fluorophore-conjugated antibodies and were analyzed by (E) flow cytometry (FACS) and (F) confocal microscopy. The results are representative of four independent experiments.

Article Snippet: Anti-HSP90 (C45G5), anti-HSP70 (D1M6J), and anti-CAP1 (D2K3J) antibodies were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Shear, Silver Staining, Comparison, Mass Spectrometry, Real-time Polymerase Chain Reaction, Western Blot, Flow Cytometry, Confocal Microscopy

Journal: eLife

Article Title: Site-specific glycosylation regulates the form and function of the intermediate filament cytoskeleton

doi: 10.7554/eLife.31807

Figure Lengend Snippet: ( A ) HeLa cells reconstituted with WT vimentin-mEmerald were infected with Chlamydia for ten hours and treated with DMSO vehicle or 50 µM 5SGlcNAc for an additional 20 hr. Cells were fixed and immunostained for MOMP (to mark individual bacteria) and cap1 (to mark the inclusion membrane), along with vimentin-mEmerald imaging. Representative images are shown. Arrows indicate extra-inclusion bacteria. ( B ) Quantification of inclusion area and number of extra-inclusion bacteria from images in ( A ). n = 60, ***p<0.001, Welch’s t-test. ( C ) Reconstituted HeLa cell lines were infected with Chlamydia for 30 hr and then fixed, stained, and imaged as in ( A ). Representative images are shown. ( D ) Quantification of inclusion area from images in ( C ). n ≥ 54, ***p<0.001, Welch’s t-test. n.s., not significant. ( E ) HeLa cells reconstituted with empty vector were infected with Chlamydia , treated with DMSO vehicle or 50 µM 5SGlcNAc, and fixed and stained as in ( A ). Inclusion size (top; n = 20, p=0.21, Student’s t-test) and extra-inclusional bacteria (bottom; n = 28, p=0.42, Student’s t-test) were quantified. n.s., not significant.

Article Snippet: A mouse antibody against MOMP (Santa Cruz, sc-57678) and a rabbit antibody against cap1 (A. Subtil, Institut Pasteur) were diluted in PBS containing 2% BSA and 0.1% Triton X-100.

Techniques: Infection, Bacteria, Membrane, Imaging, Staining, Plasmid Preparation

Journal: eLife

Article Title: Site-specific glycosylation regulates the form and function of the intermediate filament cytoskeleton

doi: 10.7554/eLife.31807

Figure Lengend Snippet: HeLa cells reconstituted with WT (n = 36), S49A (n = 40) or Y117L (n = 34) vimentin-mEmerald were infected with Chlamydia for 10 hr and treated with DMSO vehicle or 50 µM Thiamet-G for an additional 20 hr. Cells were fixed and stained for cap1 and MOMP. Inclusion area was quantified and plotted as in .

Article Snippet: A mouse antibody against MOMP (Santa Cruz, sc-57678) and a rabbit antibody against cap1 (A. Subtil, Institut Pasteur) were diluted in PBS containing 2% BSA and 0.1% Triton X-100.

Techniques: Infection, Staining