Cck 8 kit

The direct effect of the nanoparticles on untreated RAW 264.7 cells, ID8-educated RAW 264.7 cells, ID8 cells, H9C2 cardiomyocytes, and astrocytes was assessed using the cytotoxicity CCK8 Kit (Dojindo Molecular Technologies, Tubaru, Japan) following the manufacturer manual. Briefly, 5 × 10³ cells were seeded in a 96-well plate and incubated in their respective media overnight. Then, the cells were either left untreated or treated with vehicle, free DMXAA, rHDL-DPM NPs, and rHDL-DPM-DMXAA NPs for a total final volume of 100 µL. After 24 h of incubation, 10 µL of the CCK8 Kit reagent was added to the wells with incubation at 37 degrees Celsius, and the developed formazan dye color was monitored and read within 2 h at 450 nm using the Biotek Cytation 3 plate reader. The cytotoxicity results are presented as percent absorbance at 450 nm. After absorbance values are corrected with controls, the percent absorbance is calculated as follows (Equation (4)): $$\mathrm{Percent} \mathrm{absorbance} \mathrm{at} 450 \mathrm{nm}=\frac{\mathrm{Absorbance} \mathrm{at} 450 \mathrm{nm} \mathrm{recorded} \mathrm{for} \mathrm{treatment}}{\mathrm{Absorbance} \mathrm{at} 450 \mathrm{nm} \mathrm{of} \mathrm{untreated} \mathrm{control}} \times 100$$