Vitellin (Vt) and vitellogenin (Vg) profiles were analyzed in Pteromalus puparum, a pupal endoparasitoid of Pieris rapae. Non-denaturing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analyses indicated that both native Vt and Vg were likely 370 kDa in size, consisting of two subunits of approximate 206 and 165 kDa. An indirect double antibody enzyme-linked immunosorbent assay (ELISA) for monitoring hemolymph Vg and ovarian Vt levels was developed using a monoclonal antibody and a polyclonal antibody made specially against P. puparum Vt. The synthesis and uptake of Vg in this wasp was initiated immediately after adult eclosion. The hemolymph Vg and ovarian Vt reached their highest level of 0.58 and 4.51 microg per female 24 and 48 h after adult eclosion, respectively. Both Vg synthesis and uptake were in parallel with ovarian development. The Vt levels in the developing embryos decreased progressively except 12h after parasitism. Meanwhile, nine new polypeptides with sizes ranging from 59.2 to 151 kDa, possibly resulting from the limited proteolysis of Vt originally accumulated in newly laid eggs, were detected de-novo during embryonic development using Western blotting with the monoclonal antibody against Vt. These studies provide the basis for future investigation into endocrinal regulations of vitellogenesis and understanding the reproductive strategy in this wasp.