However, in this stage a substantial amount of H3K9Me1 signal was also observed in the parasite cytoplasm. For comparison, H4K5Ac showed a GSK-2018682 strictly nuclear distribution and did not co-localize with either of the PV markers at any developmental stage. Similar to previous findings, immunodetection using the H3K9Me1 antibody resulted in an extremely weak signal in total protein lysates extracted from parasite cells isolated from their host cells by saponin lysis. It has been previously shown that besides the erythrocyte membrane, saponin lysis also results in the partial disruption of the PVM. Therefore, we speculated that the vast majority H3K9Me1 could be lost from the PVM after saponin treatment. To explore this possibility, we subjected parasitized red blood cells to different concentrations of saponin, speculating that the AR7 milder saponin exposure will lead to lesser lysis of the PV. Indeed, milder lysis with 0.06%, 0.08%, and 0.10% of saponin allowed good detections of H3K9Me1 as well as the control markers: Etramp 2 and H4K5Ac. This suggests that at these concentrations saponin can lyse the erythrocyte membrane but does not affect the PV dramatically. On the other hand, the western blot signal for both H3K9Me1 and Etramp 2 was significantly reduced in Plasmodium cells lysed with higher concentration of saponin. In contrast, the signal for H4K5Ac that is localized solely in the nucleus, was unaffected by the different saponin concentrations. Taken together, H3K9Me1 and Etramp 2 antibody generated an identical pattern of the western blot signal across the set of total protein lysates generated from parasitized red blood cells with increasing concentrations of saponin. In agreement with our original hypothesis, this indicates that H3K9Me1 is localized in the PV. Importantly, the western blot signal detected by the H3K9Me1 corresponded to a protein with the predicted molecular weight of histone 3, which indicates that the IF signal yielded by this antibody is not due to possible cross-reactivity to a different P. falciparum protein.