抑制素α主动和被动免疫对哈萨克羊生殖激素含量的影响

Effects of Inhibinαon Reproductive Hormone Content in Kazakh Sheep by Active and Passive Immunity

为了研究抑制素α(INHα)主动和被动免疫对哈萨克羊生殖激素含量的影响,本试验在对抑制素α重组质粒表达菌株进行诱导表达的基础上,将经纯化、鉴定的抑制素α重组蛋白免疫接种新疆双峰骆驼,制备驼抗抑制素α多克隆抗体,并对其进行纯化,检测抗体效价,验证抗体的特异性。之后选择3~5岁、发情时间相近并处于间情期的45只成年哈萨克羊随机分为3组,分别作为抑制素α多克隆抗体免疫组(A组)、抑制素α重组蛋白免疫组(B组)及对照组(C组),每隔10 d连续进行3次加强免疫,应用ELISA法检测在绵羊繁殖活动中具有重要功能的5种生殖激素:促卵泡素(FSH)、促黄体素(LH)、孕酮素(P 4)、雌激素(E 2)、抑制素(INH),并检测血液生化指标。结果显示,IPTG的最佳诱导浓度是0.6 mmol/L,在4 h时诱导出产量较高的抑制素α包涵体蛋白,纯化后的抑制素α重组蛋白纯度较高,并具有较好的免疫原性,经进一步验证发现,制备的抑制素α抗体效价为1∶512000,该抗体可与抑制素α重组蛋白特异性结合。说明成功制备了具有免疫原性的抑制素α重组蛋白和高效价的驼抗抑制素α多克隆抗体。免疫后A组LH、P 4含量和B组FSH、LH、P 4、E 2、INH含量与C组相比差异不显著(P>0.05),而A组FSH含量和E 2含量显著高于C组(P<0.05),INH含量显著低于C组(P<0.05)。通过血液生化指标检测发现,抑制素α蛋白和抑制素α抗血清两种免疫制剂免疫后,试验动物均没有出现不良症状。说明两种抑制素α抗原均可对哈萨克羊血液生殖激素的分泌产生良好效果,相比之下抑制素α抗血清免疫效果更佳。

In order to study the effect of inhibinα(INHα)on reproductive hormone content of Kazakh sheep by active and passive immunity,the INHαprotein was expressed and purified from INHαrecombinant plasmid strain and then injected into Xinjiang camel to prepare camel anti-INHαpolyclonal antibody.The antibody titer was detected and the specificity of the antibody was verified.45 adult Kazakh sheep(3 to 5 years old),with similar estrus and interestrus,were randomly divided into 3 groups,which were respectively used as the INHαpolyclonal antibody immunization group(group A),the INHαrecombinant protein immunization group(group B)and the control group(group C).These sheep were immuned three times every ten days.The ELISA method was used to detect five reproductive hormones that had important functions in sheep reproduction:follicle stimulating hormone(FSH),luteinizing hormone(LH),pregnancy ketone(P 4),estrogen(E 2),INH(inhibin).Blood biochemical indicators were detected.The results showed that the optimal induction concentration of IPTG was 0.6 mmol/L,and a higher yield of INHαinclusion body protein was induced at 4 h.The INHαprotein was highly purified and had good immunogenicity.The prepared INHαantibody had a titer of 1∶512000,and the antibody could specifically bind to INHαprotein.It showed that an immunogenic INHαprotein was successfully prepared and after further verification.The camel anti-INHαpolyclonal antibody with high antibody titer was successfully prepared.After immunization,the LH,P 4 content of group A and FSH,LH,P 4,E 2,INH content of group B compared with group C were not significantly different(P>0.05),while the FSH and E 2 contents of group A were significantly higher than those of group C(P<0.05),the INH content was significantly lower than that of group C(P<0.05).Through the detection of blood biochemical indicators,it was found that the experimental animals had no adverse symptoms after the immunization of the two immune preparations of INHαprotein and INHαantiserum.The results showed that both INHαantigens could have a good effect on the secretion of blood reproductive hormones in Kazakh sheep,and the INHαantiserum had a better immune effect.