GM-CSF和FliC作为新型复合生物佐剂对猪圆环病毒疫苗的免疫增强效果

Immune enhancement effect of GM-CSF and FliC as a new compound biological adjuvant on porcine circovirus vaccine

研究成功制备猪粒细胞-巨噬细胞集落刺激因子(GM-CSF)和鼠伤寒沙门氏菌鞭毛蛋白(FliC),与猪圆环病毒疫苗共同作用形成高效佐剂效应,为进一步应用复合生物佐剂奠定基础。GM-CSF在37℃下经终浓度0.25 mmol·L-1IPTG诱导4 h大瓶培养,以包涵体形式表达,变性、复性后,经Ni-NTA柱亲和层析,最终得到GM-CSF蛋白,大小约13 ku。FliC在25℃下经终浓度0.25 mmol·L-1IPTG诱导6 h发酵培养,以可溶形式表达,经纯化、蛋白酶酶切后,最终得到约46 ku FliC蛋白。将制备蛋白与猪圆环病毒疫苗共同注射小鼠,经ELISA检测发现GM-CSF、FliC均可单独作为生物佐剂有效提高抗体效价,促进免疫。与单独佐剂组相比,复合佐剂组起效更迅速,作用时间更持久,作为一种新型复合生物佐剂应用前景良好。

The study successfully prepared GM-CSF(pig granulocyte-macrophage colony stimulating factor)and FliC(Salmonella typhimurium flagella)protein,got effective adjuvant effect of them with porcine circovirus vaccine,and laid a experimental foundation for the further application of compound biological adjuvant.GM-CSF at 37℃under the final concentration of 0.25 mmol·L^-1 IPTG for 4 h was expressed in the form of inclusion body.After denaturing and refolding,affinity chromatography was performed through NI-NTA column to obtain the GM-CSF protein,which was about 13 ku in size.FliC at 25℃under the final concentration of 0.25 mmol·L^-1 IPTG for 6 h fermentation cultivation was expressed in soluble form.After purification and protease digestion,the FliC protein was approximately 46 ku in size.The prepared proteins were injected into mice with the porcine circovirus vaccine.It was found by ELISA that GM-CSF and FliC alone could effectively increase the antibody titer as a biological adjuvant,both could promote humoral immunity.Compared with the single adjuvant group,the compound biological adjuvant group worked more rapidly and for a longer time,which was a new type of compound biological adjuvant with great potential application prospect.