High speed ratiometric calcium imaging
Human sperm cell labelled with 2 single wavelength calcium probes (Fura Red and Fluo-4) and treated with NPEGTA (caged calcium). We used Cairn's beam spliter (Optosplit II) to project emissions (515nm and 650nm) from both fluorophores to adjacent areas of the same camera and generate high speed (60Hz) ratiometric videos illustrating intracellular calcium concentration over time on tethered human spermatozoa to study flagellar beat. Cells were exposed to a localised UV flash to trigger a calcium bolus mid flagellum to reveal calcium signal propagation.
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