HPMVEC were grown to confluence on glass cover slips in 12 well plates. The cells were washed three times with serum-free medium and kept for 2 h at 37°C in medium containing 1% FBS before agonist treatment. After thrombin, heparin, ODSH treatments, cells were washed three times with HBSS and fixed with 1% paraformaldehyde for 15 min at room temperature (RT). Cells were then blocked with 5% BSA at 4°C for 60 min prior to incubation with anti-PAR-1 rabbit polyclonal IgG (1:40) (Santa Cruz Biotechnology, Santa Cruz, CA, USA). The anti-PAR-1 antibodies were detected after incubation of the cells with rhodamine-labeled goat anti-mouse antibodies (1:400) for 30 min at 4°C. Cell surface fluorescence was visualized and photographed using Zeiss Axio Observer imaging system.
Axio observer imaging system
Manufactured by Zeiss
The Axio Observer imaging system is a high-performance microscope designed for advanced imaging applications. It features a modular design, allowing for customization to meet specific research needs. The Axio Observer provides precise control over illumination, optics, and image acquisition to enable detailed analysis of samples.
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Lab products found in correlation
2 protocols using axio observer imaging system
1
Quantification of PAR-1 Expression in HPMVEC
2
Immunofluorescence Staining of USP7 and TRAP
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Immunofluorescence staining was performed as detailed by Xie et al. [19 (link)]. As mentioned above, the slices were dewaxed with xylene and ethanol. The cells were then fixed with 4% paraformaldehyde and incubated with 0.1% Triton X-100 at room temperature for 10 min. After blocking with goat serum, the cells were incubated with anti-USP7 (Abcam 264,422) and anti-TRAP (Abcam 191,406) antibodies overnight at 4 °C. The appropriate secondary antibody was subsequently added, and the cells were incubated at room temperature for 1 h. The sections were then restained with 4′,6-diamino-2-phenylindole (DAPI) to label the nucleus. After application of anti-fading installation medium (P0126 Magi Beyotime), the slides were covered with cover slips. An image of the sample was captured using an Axio Observer imaging system (Carl Zeiss).
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