2.4. Measuring the reactive oxygen species

Total ROS was measured by staining the cells using the 2',7'-dichlorofluorescin diacetate (DCFDA) cellular ROS detection assay kit (Beyotime, China). Blood samples were collected and incubated with 2,7-dichlorodihydrofluoresceine diacetate (DCFH-DA) at 37°C for 20 min. Thereafter, the cells were washed with PBS and dichlorofluorescein (DCF) fluorescence determined using a fluorescence microplate reader (Safire2 Tecan, Mannedorf, Switzerland). The excitation filter was set at 485 nm whereas the emission filter was set at 528 nm.