论文部分内容阅读
哺乳动物卵透明带(ZP)是避孕疫苗可能的候选抗原之一。研究结果表明人ZPA重组表达蛋白在兔体内诱导了可抑制人体外受精的抗血清。本研究旨在确定该重组蛋白是否也能在一非人灵长类动物中产生可阻断受精的抗血清。 方法 免疫原使用了去除信号肽的猪ZPA氨基端198和人ZPA氨基端206个氨基酸残基肽,它们均山含各重组pET21b质粒的大肠杆菌(BL21)表达,然后被分别纯化用作抗原、帽猴的免疫采用完全福氏佐剂和单次为500μg抗原的背部两位点肌肉注射方式,在每次间隔1个月的4次加强免疫后收集抗血清。关于抗血清效价的鉴定,抗体生成的测定采用人卵母细胞免疫荧光染色法和酶联免疫吸附试验(ELISA)法,阻断受精试验使用人卵母细胞和精子。
Mammalian zona pellucida (ZP) is one of the potential candidate antigens for contraception. The results showed that human ZPA recombinant protein induced in rabbits can inhibit human antiserum in vitro antiserum. The aim of this study was to determine if this recombinant protein could also produce antisera to block fertilization in a non-human primate. Methods The immunogen used the porcine ZPA amino terminus 198 with the signal peptide removed and the amino acid residue 206 amino acid residues of the human ZPA both expressed in E. coli BL21 containing the respective recombinant pET21b plasmids and was then purified separately for use as antigen, The immunization of the capitatum was performed by means of intramuscular injection of both Freund’s adjuvant and a single dose of 500 μg of antigen on both sides of the back and collecting the antiserum after four boosts at intervals of one month. For the identification of antiserum titer, human oocyte immunostaining and enzyme-linked immunosorbent assay (ELISA) were used for the determination of antibody production, and human oocytes and sperm were blocked using a fertilization test.