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For the four-step gradient, layer 1 ml of each of 4%, 5.5%, 7% and 10% (w/v) iodixanol and layer the cell suspension on top OR for the two step gradient layer an equal volume of 9.6% (w/v) iodixanol on top of the cell suspension in 21.6% (w/v) iodixanol (see Notes 4 and 5)

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Hydrogen sulfide (H 2 S), a gaseous signaling molecule produced by some cells within the body, acts as a vasodilator (relaxes blood vessels) and thus may have cardioprotective properties (51, 52)