Iron deficiency, one of the most important nutritional problems in the world, can be caused not only by foods deficient in iron but also by poor availability of dietary iron. Iron food fortification in combination with highly available iron from supplements could effectively reduce this deficiency. The aim of this study was to examine the iron availability from iron-fortified spirulina. We have used an in vitro digestion/Caco-2 cell culture system to measure iron spirulina availability and made a comparison with those of beef, yeast, wheat floor, and iron sulfate plus ascorbic acid as a reference. Iron availability was assessed by ferritin formation in Caco-2 cells exposed to digests containing the same amount of iron. Our results demonstrate a 27% higher ferritin formation from beef and spirulina digests than from digests of yeast and wheat flour. When iron availability was expressed per microgram of iron used in each digest, a 6.5-fold increase appeared using spirulina digest in comparison with meat. In view of this observed high iron availability from spirulina, we conclude that spirulina could represent an adequate source of iron.