For flow cytometry analysis, monolayers of exosome-treated HUVECs and HPMECs were detached from culture flasks by incubation on ice before being washed twice in PBS. Detached cells were resuspended in 200 µL of PBS containing 1% BSA (Sigma-Aldrich) and incubated for 30 min at 4 °C with the following antibodies: PE-conjugated anti-CD144, PE Mouse IgG1, and κ Isotype Control (BD Biosciences). After washing, the cells were analyzed using a FACScan laser flow cytometer, BD FACS AriaII (BD Biosciences). Mean channel fluorescence was expressed as FACS arbitrary values. Finally, the cells were gated using forward vs. side scatter to exclude dead cells and debris.
Facscan laser
Manufactured by BD
Sourced in United States
The FACScan laser is a flow cytometry instrument designed for cell analysis and sorting. It utilizes a laser light source to excite fluorescent labels within cells, enabling the detection and quantification of various cellular properties and characteristics. The FACScan laser provides core functionality for flow cytometry applications without interpretation or extrapolation on intended use.
Automatically generated - may contain errors
Lab products found in correlation
Propidium iodide, by Merck Group (2 mentions)
Pe mouse igg1, by BD (1 mentions)
κ isotype control, by BD (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Facsaria 2, by BD (1 mentions)
Cellquest software, by BD (1 mentions)
Cellquest, by BD (1 mentions)
Cellquest version 3, by BD (1 mentions)
6 protocols using facscan laser
1
Exosome-Induced Endothelial Cell Analysis
2
Flow Cytometric Analysis of Apoptosis and Cell Cycle
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For cell apoptosis assay, the HS-5 cells were cultured then treated as described in groups of experiment. After 48h treatment, cells were harvested and centrifuged at 1000r/min for 5min.Subsequently, cells were resuspended with 500µ L PBS solution for each tube. Cell apoptosis was detected by the flow cytometry. For cell cycle assay, the HS-5 cells were cultured then treated as described in groups of experiment. After 48h treatment, cells were harvested and fixed with pre-cooled 75% ethanol at 4°C for at least 5h. After centrifugation, cells were incubated with propidium iodide (PI) and RNase A at 37°C for 30 min in dark. Cell cycle was detected by the flow cytometry. The apoptosis and cell cycle were analyzed on a FAC-Scan laser flow cytometry (BD Biosciences, New Jersey, the USA). The data were processed by Cell Quest software (BD Biosciences, New Jersey, the USA).
3
Cell Cycle Analysis by Flow Cytometry
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Cell cycle population was determined by flow cytometry as follows. After exposing to HMDB for 0, 6, 12, and 24 h, HeLa cells were washed twice with PBS, and then fixed in 70% ethanol for additional 2 h at −20 °C. Following fixation, cells were washed with PBS again, incubated with 1 mL of PBS containing 0.5 μg/mL RNase A and 0.5% Triton X-100 for 30 min at 37 °C. Then the cells were stained with 50 μg/mL propidium iodide (PI). The stained cells were estimated by a FACScan laser flow cytometer equipped with Cell Quest software (Becton Dickinson, San Jose, CA, USA).
4
Cell Cycle Analysis of Synchronized Cells
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Cells were washed with PBS three times 24 h after plating and incubated with media containing 0.04% fetal calf serum (FCS) for an additional 24 h. Under these conditions, cells are arrested in G0/G1 phase based on flow cytometry analysis as reported in our previous study [18 (link)]. Next, synchronized cells (cultured in 0.04% FCS) were challenged with the addition of media containing 10% FCS. Apiole- and PBS-treated groups were assessed via flow cytometry analysis to determine cell cycle distribution. Cells were stained with propidium iodide (50 μg/ml) (Sigma Chemical Co., St. Louis, MO, USA), and DNA content was assessed using a FACScan laser flow cytometry analysis system (Becton-Dickinson, San Jose, CA, USA), with 15,000 events being analyzed for each sample.
5
Apoptosis Analysis of HeLa Cells Treated with Fungal Polysaccharides
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Seeded HeLa cells were incubated with medium containing inhibitory concentrations of total polysaccharide extracted from O. erinaceus overnight. Then, cells were washed with PBS and suspended with PI solution containing 0.1% sodium citrate plus 0.1% Triton X100 at 37°C for 30 min and then placed at 4°C in the dark for 10 min and the apoptotic cells were evaluated using a FACScan laser flow cytometer (FACS Calibur, Becton Dickinson, USA).
6
Cell Cycle Analysis of VSMC by Flow Cytometry
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The quantification of cell cycle distribution was examined using a FACScan laser flow cytometer (Becton Dickinson, San Jose, CA, USA). The VSMC was treated with TNF-α (10 ng/mL) in the absence or presence various concentrations (0.2 and 0.5 mg/mL) of HLP for 24 h; collected, rinsed with PBS twice; fixed in 70% ethanol at –20 °C overnight; and then stained with propidium iodide (PI) solution (20 μg/mL of PI, 20 μg/mL of RNase A, and 0.1% Triton X-100; all chemicals from Sigma-Aldrich, St Louis, MO, USA) for 20 min in the dark at room temperature. Each phase of cell cycle was presented as the cell number versus the DNA content as indicated by the intensity of fluorescence, and gated into subG1, G0/G1, S, and G2/M phases with CELLQuest Version 3.3 software (Becton Dickinson, San Jose, CA, USA).
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