Sickle cell disease (SCD) is one of the commonest severe inherited disorders. All SCD patients have the abnormal haemoglobin HbS in their red blood cells (RBCs) instead of the normal adult HbA. Deoxygenated HbS polymerises and distorts RBCs into bizarre shapes. The resulting pathology is extensive, with anaemia and vaso-occlusive episodes of pain, organ damage and mortality. Early diagnosis is critical to manage the severest complications of SCD [3]. RBCs from SCD patients have a high cation permeability causing solute loss, shrinkage and raised [HbS]. As the polymerisation lag time is inversely proportional to a very high power of [HbS], modest shrinkage is sufficient to encourage greatly HbS polymerisation and thereby contribute to disease. Several pathways participate in the increased permeability [2]: KCl cotransport, the Gardos or Ca2+-activated K+ channel and a deoxygenation-induced non-selective cation pathway (Psickle). Psickle has a central role mediating Ca2+ entry, activating the Gardos channel and causing other sequelae such as phosphatidylserine exposure. Although the principle defect is increased cation permeability, deoxygenated RBCs from SCD patients have been shown to haemolyse in isosmotic non-electrolyte solutions [1]. Lysis is accompanied by entry of non-electrolyte and a number of features suggestive of Psickle involvement [1]. As Psickle is only activated in RBCs from SCD patients, this phenomenon may be used to design a simple diagnostic test for SCD which might be prognostically useful. Here we further characterise this assay. RBCs (from discarded samples) were washed in MOPS-buffered saline (in mM: NaCl 145, glucose 5, MOPS 10, pH 7.4, 290±5mOsm.kg-1, 37oC unless otherwise stated) and re-suspended in isosmotic sucrose solution (sucrose replacing NaCl). At pH 7.4, RBCs from HbSS and HbSC SCD patients lysed on deoxygenation; those from HbAA and HbAS individuals did not (Fig 1a). Ouabain (100µM), bumetanide (10µM), or raising or lowering [sucrose], had minimal effect on lysis. Clotrimazole (10µM) or EGTA (2mM) modestly, but repeatably, increased lysis. Deoxygenation in the presence of o-vanillin (5mM) to prevent HbS polymerisation abrogated haemolysis. Effect of pH was studied. At pH 6, RBCs from SCD patients lysed even when oxygenated, an effect also prevented by o-vanillin (Fig 1b). Findings show that haemolysis in isosmotic non-electrolyte solutions appears exclusive to RBCs from SCD patients. Gardos channel activation and KCl loss reduces lysis by only a small extent. Hypo- or hypertonicity to alter RBC volume or chemical driving force for sucrose entry is without effect. The inhibitory effect of o-vanillin is consistent with HbS polymerisation being a pre-requisite for lysis. The ability to use oxygenated solutions at low pH will facilitate development of the test.