After removal of the surrounding connective tissue, femurs were dissected from the three groups of mice. Femurs were clipped in DPBS digested with 2.5 mg/mL collagenase A (Biosharp, Hefei, China) and enzymatically with 1 unit/mL dispersible enzyme I (Biosharp, Hefei, China) and kept under gentle agitation at 37°C for 15 min. The obtained cell suspension was filtered using a 40-um cell strainer (Biosharp, Hefei, China) and washed with DPBS. After washing, the cells were incubated with FITC-conjugated EMCN antibody (Santa Cruz Biotechnology, USA) and APC-conjugated CD31 antibody (BioLegend, San Diego, CA) for 30 min at room temperature. Thereafter, the cells were resuspended in DPBS. All samples were analyzed using the CytoFLEX LX Flow Cytometry System (CytoFLEX, Backman Counter, CA, USA) and CytExpert 2.4 Software.
Cytoflex lx flow cytometry system
Manufactured by Beckman Coulter
Sourced in United States
The CytoFLEX LX Flow Cytometry System is a multicolor flow cytometer designed for high-performance cell analysis and sorting. It features advanced optics and electronics to enable the simultaneous detection of up to 21 parameters.
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Lab products found in correlation
2 protocols using cytoflex lx flow cytometry system
1
Isolation and Characterization of Endothelial Cells
2
Comprehensive Flow Cytometry Analysis
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Flow cytometry was performed as described in previously research (Ji et al., 2019 (link)). Briefly, Single cell suspensions were stained with the indicated antibodies diluted by PBS supplemented with 2% FBS and 0.5% BSA for surface markers. For Foxp3 and ROR‐γt staining, the cells were stained for surface markers such as anti‐CD45 and anti‐CD4, followed by fixation and permeabilization with fixation and permeabilization buffer (#88–8,824‐00; Thermo Fisher Scientific, America) at room temperature for 30 min. After washed with PBS for three the cells were then stained with anti‐Foxp3 and anti‐ROR‐γt antibodies as instructed. All samples were detected and analyzed by CytoFLEX LX Flow Cytometry System.
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