Figure 3. Western blot analysis to determine the onset of vitellogenin synthesis in honeybee queen and worker pupae. SDS-PAGE (7,5%) was performed with 1mg of hemolymph proteins. After blotting to a PVDF membrane vitellogenin was detected by a rabbit antiserum raised against honeybee egg proteins in combination with the PAP-DAB detection system. A) queen and B) worker pupae treated as brown-eyed pupae (Pb) with 10 mg juvenile hormone dissolved in acetone (treated), or with acetone (controls). C) Hemolymph from worker pupae of Apis mellifera treated, in Pb phase, first with a single injection of 10 mg of actinomycin D, and 1h later, with 10 mg of juvenile hormone-III. Controls received 10 mg of juvenile hormone-III. Pbl, Pbm and Pbd: brown-eyed pupae showing, light, medium, or dark pigmented cuticle, respectively. Pbm I , Pbm II and Pbm III are progressive ages within Pbm phase. Arrows indicate the 180 kDa vitellogenin band.