Our group attempted the first DV pH measurements for living intra-erythrocytic parasites under physiologic perfusion using the pH probe acridine orange (AO) and novel SCP [71, 82] . Our initial hypothesis was that, owing to the dibasic nature of CQ, even subtle increases in DV pH would significantly lower DV concentrations of CQ and thus cause CQ resistance. Therefore, relative to CQS, CQR parasites might show slightly elevated DV pH or different DV pH behavior upon addition of CQ [83] , or perhaps both. We and others have actually found that mutant PfCRT in CQR parasites causes more acidic (lower) DV pH. The initial AO data in support of this conclusion generated controversy (as most new technologies tend to do), but a number of different, complementary methods from several labs were subsequently developed and strongly supported the initial AO conclusions [21, 22, 74, 76, 79] .