MEASUREMENT OF pH OF THE DIGESTIVE VACUOLE ISOLATED FROM THE PLASMODIUM FALCIPARUM-INFECTED ERYTHROCYTE BY DIGITONIN PERMEABILIZATION

The acidic pH of the digestive vacuole (DV) of P. falciparum is thought to play a major role in the mechanisms of digestion of host erythrocyte hemoglobin, detoxification of heme, and chloroquine (CQ) action and resistance. However, a definitive measurement of the DV pH has been technically difficult. A previous study by Abu Bakar (2015) measured the DV pH by using fluorescein isothiocyanate-dextran (FITC-dextran), a ratiometric pH indicator loaded into the malaria parasite’s DV that had been isolated using saponin (DV_sap). To validate the FITC-dextran response to the DV_sap pH, pH of the DV isolated by digitonin permeabilization (DV_digi) was measured. The DV_digi pH of the CQ-sensitive parasite (D10) was 5.66 ± 0.07 that is approximately similar to that of the DV_sap pH (5.27 ± 0.03) estimated using the same probe. The DV_digi pH of the CQ-resistant parasite (Dd2) (5.62 ± 0.12) was not significantly different from the CQ-sensitive parasite’s DV_digi pH (P> 0.3). Re-acidification of the DV_digi was also observed upon the addition of 2 mM ATP to the ATP-depleted medium. These data validate the use of FITC-dextran for quantitative DV pH analysis of the parasite isolated using either digitonin or saponin.