All serum of seroresistant syphilitic patients and healthy controls were collected by sterile tubes containing separating glue with centrifuging at 900 g for 5 min and stored at −80℃ for subsequent use. The concentrations of serum chemokines were quantitatively measured by Cytometric Bead Array (CBA) Human Chemokine Kit(552990)comprising microbeads coupled with monoclonal antibodies (MoAb) against CXCL8, CCL2, CXCL9, CCL5, and CXCL10. The levels of the corresponding chemokines were acquired using a FACS Verse flow cytometer (Beckman Coulter, Inc.) according to the manufacturer's instructions (Beckman Coulter, Inc.) and analyzed by BD FCAP Array 3.0 software (Becton Dickinson and Company).
Facs verse flow cytometer
Manufactured by Beckman Coulter
Sourced in United States
The FACS Verse flow cytometer is an analytical instrument designed for cell analysis. It utilizes the principles of flow cytometry to rapidly identify and quantify various cell types within a sample. The core function of the FACS Verse is to provide precise measurements of physical and biochemical characteristics of cells passing through a laser-based detection system.
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4 protocols using facs verse flow cytometer
1
Quantifying Chemokine Levels in Syphilis
2
Apoptosis Analysis by Flow Cytometry
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Flow cytometry analysis was performed to identify apoptosis by Annexin V‐FITC/Propidium Iodide apoptosis detection kit (Cell Signaling Technology). After treatment, cells (1 × 105 cells/30 mm dish) were harvested and stained with Annexin V‐FITC/PI according to the manufacturer's procedure. The cells were analyzed by FACS Verse flow cytometer (Beckman Coulter, Brea, CA, USA).
3
Annexin V Staining of Mouse Erythrocytes
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Mouse blood cells were washed twice using Ringer’s solution supplemented with 5 mM calcium chloride. To detect FITC annexin V-positive cells, the erythrocytes were suspended in an annexin-binding buffer (BD Pharmingen, San Diego, United States) with FITC annexin V (1:200 dilution, BD Pharmingen, San Diego, United States) and incubated for 15 min at room temperature. Well mixing was performed by pipetting. Finally, erythrocytes were diluted five times in the annexin-binding buffer before analysis in the FACSVerse flow cytometer (Beckman Coulter, CytoFlex S, United States) at an excitation wavelength of 488 nm (blue laser) and emission wavelength of 530 nm.
4
Cell Cycle Analysis by Flow Cytometry
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The cells were cultured for 48 h following transfection, then digested with 0.25% non-EDTA trypsin, followed by centrifugation at 300 × g, at room temperature for 5 min to harvest the cells. The cells were then resuspended in 100 ml PBS, and fixed by slowly adding 700 ml pre-cooled 80% ethanol to reach a final concentration of 70%, following incubation at 4°C for 4 h before centrifugation at 300 × g for 5 min, at 4°C. Then, RNase (100 ml; 50 µg/ml) was added, and the cells were placed in a water bath at 37°C for 30 min to permeabilize the cells. Finally, PI solution (400 µl; 50 µg/ml) was added, and the cells were stained at 4°C for 30 min in the dark. The cell cycle distribution was evaluated using FACSVerse™ flow cytometer (Beckman Coulter, Inc.) and the data were analyzed using FlowJo v10 software (FlowJo LLC). The experiments were performed independently 3 times.
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