For GFP-IP, 1 mL of supernatant was incubated at 4 °C for 3 4 h with 25 μL of GFP-Trap_A beads (Chromotek, Planegg-Martinsried, Germany).
The concentrations of these cytokines were calculated according to standard curves, and the obtained results are shown as the amount (pg) of IL-1β, IL-6, TNF- α and IL-10 per mL of supernatant.
To initiate reaction, 0.1 mL of supernatant was added.
Then, 1.8 mL of supernatant was removed cautiously, and the cells were resuspended by vortexing.
After centrifugation, 2.0 mL of supernatant and 2.0 mL of 0.25 % TBA reagent were mixed.
After the centrifugation, 0.3 mL of supernatant was carefully removed using pipette and ICP-MS analyzed.
Two ml of 50%% (w/v) cold trichloroacetic acid (TCA) were added to 10 ml of supernatant.
0.1 ml of supernatant was taken and 5 ml of dye mixture was added and used for sample analysis.
After centrifugation, 1.0 mL of supernatant and 2.0 mL of 0.25 % of thiobarbituric acid (TBA) reagent were mixed.
After filtration of the mixture, 5 mL of supernatant solutions were analyzed by liquid scintillation spectrometry (LSC) instrument.
Then, 0.5 mL of supernatant was mixed with 0.5 mL of potassium phosphate buffer (10 mM, pH 7.0).
