Red Blood Cell Sickling During Oxygen Cycles in a Microdroplet Device

We have developed a novel microfluidic device to study repetitive sickling on individual red blood cells by replicating the physiological oxygen cycling of the vascular circulatory system (Abbyad et al., Lab Chip, 2011, 11, 813). A small number of red blood cells from sickle cell patients are encapsulated in an array of aqueous microdroplets. These microdroplets are anchored and arranged in a 2-dimensional array against the flow of the carrier oil. Precise spatial and temporal changes in oxygen concentration are obtained through gas exchange with the inert oil flowing outside the droplets. By oscillating the oxygen concentration, cycles of sickling and desickling of individual red blood cells are observed in real-time. Polarization microscopy allows for the sensitive detection of intracellular hemoglobin fibers. We observed small residual intracellular hemoglobin fibers that remain even in oxygenated conditions. Since the content of droplets in the array can be controlled, active molecules at different concentrations as well as control droplets can be measured side-by-side as they are exposed to the same environmental conditions. This was used to measure cell sickling in the presence and the absence of the anti-sickling agent glyceraldehyde. The cumulative impact of repeated sickling, such as membrane damage and cell dehydration, is believed to be central to disease pathology. We are now studying phosphatidylserine outer leaflet externalization and cell dehydration as a function of deoxygenation cycle.