We investigated the role of 20-hydroxyecdysone (20E) on the ovarian maturation and protein levels and pattern in adult females of the spider Tegenaria atrica. In unmated females, the ovaries did not initiate vitellogenesis and levels of proteins in the hemolymph and ovaries were low. Unmated females treated with 20E showed an increase in the total amount of proteins in their hemolymph and ovaries, comparable with that in untreated mated females. Vitellogenesis was observable histologically in 20E treated unmated females as in untreated mated ones. The electrophoretic analysis of total proteins revealed a 47 kDa band expressed in the vitellogenic ovaries and hemolymph of 20E treated unmated females and untreated mated ones as in newly laid eggs. We hypothesize that this 47 kDa protein could belong to the vitellogenin-vitellin complex. We describe in this paper, for the first time in a spider, the partial characterization of putative yolk protein.