His tag mouse mcab

Western blot analysis and qRT-PCR were conducted as in Lee et al.[15 (link)]. His-Tag antibody (HRP-conlugated 6*His, His-Tag Mouse McAb, Proteintech) was diluted 1: 5000 for western blot. The qRT-PCR primer pairs used in this study are listed in S1 Table.

PLY could bind to the cholesterol membranes of cells to causing cytolysis via self- binding oligomers formation [8 (link)]. In our study, oligomerization analysis was performed to explore whether shionone could disturb the oligomers formation to neutralize the activiy of PLY. PLY (0.48 μM) was coincubated with different concentrations of shionone (2, 4, 8 μg/mL) in PBS at 37 °C for 1 h. Afterward, 5 × SDS loading buffer without beta-mercaptoethanol was added and incubated for 10 min at 55 °C. Samples were separated by 6% SDS-PAGE and transferred to PVDF membranes. Moreover, PVDF membranes were incubated with the His-Tag Mouse McAb (Proteintech, Rosemont, IL, USA) for 1 h and analyzed with a visualizer after washing 3 times.