Fig. 4. Maternal hb expression in S. americana. Northern analysis, in situ hybridization and immunostaining reveal that hb is expressed during oogenesis. One transcript of 4.2 kb is expressed in the L. migratoria ovary (A), with an additional transcript of 6.0 kb appearing during embryogenesis. (B-E) In situ hybridization results using antisense (B-D) and sense (E) hb probes on S. americana ovarioles. Expression begins within the oocyte, as it leaves the germarium (B, arrow): there is no detectable expression in the germarium (left of arrow) or the somatic cells that surround the oocyte. By mid-oogenesis, hb transcript is abundant (C), but fades to background levels in older oocytes (D). (E) Control hybridization with a sense probe of oocytes at about the same stage as those seen in C. (F-J) Immunostaining of S. americana ovaries with monoclonal antibody PP 7C11. In F, the almost completed egg (*) is rotated relative to the rest of the ovariole and the presence of a vitelline membrane around it prevents the penetration of antibodies; (g) indicates the position of the germarium. Hb protein is expressed at high levels within developing oocytes and is localized to the germinal vesicle (oocyte nucleus). Translation begins around the time that transcript can first be detected (G), and continues through mid-oogenesis (H) until the germinal vesicle moves posteriorly in late oogenesis (arrowheads in F). As the nucleus moves posteriorly, Hb protein is still nuclear (I), but seems to exit the posteriorly located nucleus about the same time that the egg undergoes a phase of rapid expansion as it fills with yolk (J).