Figure 6
Effect of glucose oxidase on annexin V staining of HuH7 cells. (A) Flow cytometry. Following glucose oxidase treatment, cells were harvested, washed and stained with annexin V and propidium iodide. Glucose oxidase increased the proportion of cells staining with annexin only and with both annexin and PI. (B) Immunocytochemistry. Cells were seeded at 2 × 104 per well in eight-well chambered slides, incubated overnight and treated for 2 h with 75 mU ml−1 glucose oxidase. The medium was then harvested and cytospun on to charged slides at 300 g for 3 min. Cytospins were stained with annexin V and propidium iodide and visualised using fluorescence microscopy. The majority of cytospun cells had a necrotic phenotype. However, some showed features of apoptosis including loss of cytoplasm, blebbing and nuclear condensation (top cell). Almost all cells co-stained with annexin and PI suggesting any apoptotic cells had progressed to lose membrane integrity.
