title: [Damage of seminiferous tubules in mouse testis caused by active immunization of actin-like protein 7a].
abstract: OBJECTIVE
  To obtain recombinant sperm-protein actin-like protein 7a (ACTL7a) and detect the damage seminiferous tubules in mouse testis caused by anti-sperm antibodies generated by purified ACTL7a active immunization.
  
  
  METHODS
  The recombinant expression plasmid pET30a-ACTL7a was constructed and then transformed into E. coli Rosseta (DE3). The protein expression was induced by isopropyl β-D-1-thiogalactopyranoside (IPTG), and the protein was purified by nickel ions chelating resin. Finally, the protein was separated by sodium dodecyl sulfate poly-acrylamide gel electrophoresis (SDS-PAGE) and harvested by excising the gel containing target. ICR (Institute of Cancer Research) mice were then immunized using purified ACTL7a protein. The antibody titers were determined by ELISA and the development of seminiferous tubules after active immunization was stained by PAS staining.
  
  
  RESULTS
  Induced by IPTG, the target protein ACTL7a was expressed in E. coli. After purification, it was used to immunize the ICR mice. As shown by PAS staining, spermatid expulsion, pyknotic cells, absence of germ cells, and germ cells degenerated were seen in the seminiferous tubules in the immunized testes.
  
  
  CONCLUSIONS
  The ACTL7a prokaryotic expression vector was successfully constructed. High-purity target protein was obtained after induction and purification. After the active immunization with the target protein, the seminiferous tubules in the mouse testes will be severely damaged.
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
authors.forename: Jun
authors.initials: J
authors.lastname: Fu
authors.affiliation: Department of Reproductive Physiology, National Research Institute for Family Planning Beijing, Beijing 100081, China.
authors.forename: Shu-dong
authors.initials: SD
authors.lastname: Zong
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
authors.forename: Shi-ying
authors.initials: SY
authors.lastname: Miao
authors.affiliation: State Key Laboratory of Medical Molecular Biology, Department of Biochemistry and Molecular Biology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Song
mesh_terms: D000199:Actins / Q000009:adverse effects* / Q000378:metabolism; D000818:Animals; D000906:Antibodies; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D021381:Protein Transport; D011994:Recombinant Proteins; D012671:Seminiferous Tubules / Q000378:metabolism / Q000473:pathology*; D013094:Spermatozoa; D013737:Testis / Q000378:metabolism; D014611:Vaccination / Q000009:adverse effects*
chemical_list: D000199:Actins; C483118:Actl7a protein, mouse; D000906:Antibodies; D011994:Recombinant Proteins