2.10. Biocompatibility studies

Biocompatibility of Co₃O₄-NPs and MgO-NPs was evaluated against isolated human red blood cells (hRBCs), employing previously used methodology (Khalil et al. 2017). In the experiment, 2 mL of healthy blood was collected in EDTA tube, with an informed consent, from a volunteer. To isolate the erythrocytes, the collected blood was centrifuged at 13000 rpm for 10 min. After centrifugation, the supernatant was discarded and the pellet was washed thrice with PBS. After thorough washing, the pellet was mixed with 9.8 mL of PBS (pH: 7.2), with gentle shacking to prepare the PBS-erythrocyte suspension. Tested samples with various concentration and erythrocyte suspension were incubated for an hour in Eppendorf tubes at 35 °C, followed at 12,000 rpm for 8 min. From the centrifuged eppendorf tubes, 200 μL of the supernatant was transferred into corresponding well in a 96-well plate. Using a microplate reader, absorptions were measured for haemoglobin release at 540 nm. Triton X-100 (0.5) and Dimethyl Sulfoxide were employed as positive and negative controls. % hemolysis was calculated as

Where Abs, Abnc and Abpc denotes absorbance of sample, negative control and positive control respectively.