The systematics of the Asplenium seelosii complex have been debated for a long time. This complex includes strictly rupicolous plants that live on limestone cliffs mainly in mountains of southwest Europe: the Alps, the Pyrenees, and several mountain ranges of the Eastern Iberian Peninsula. The disjunct distribution of the populations and several morphological characters, i.e., leaf indumentum and the structure of the perispore, have been used to distinguish species and subspecies. The goal of this study was to evaluate the different systematic treatments of this complex by means of isozyme electrophoresis. Seventeen populations throughout the range of the complex were studied, and 15 enzymatic systems were assayed. There was no within population genetic variation and genetic identity between populations varied widely. Analysis of isozymic data clearly differentiated two groups corresponding to the species proposed, Asplenium seelosii and A. celtibericum, but these data do not support the recognition of subspecies in this complex.