STAg联合CpG ODN和CT鼻内免疫小鼠诱导的特异性抗体应答和脾淋巴细胞的特异性增殖反应

The specific antibody response and splenic lymphocyte proliferative response of mice following intranasal immunization with STAg plus CpG ODN and CT

目的观察可溶性速殖子抗原(STAg)联合CpG寡核苷酸(CpG ODN)和/或霍乱毒素(CT)鼻内免疫BALB/c小鼠后血清IgG、肠冲洗液IgA抗体水平及脾淋巴细胞体外特异性增殖反应,探讨CpG ODN和CT的佐剂效应。方法6周龄BALB/c小鼠60只,随机分为5组,分别用20μl PBS(PBS组)、20μg STAg(抗原组)、20μg STAg+10μgCpG ODN(CpG佐剂组)、20μg STAg+1μg CT(CT佐剂组)、20μg STAg+1μg CT+10μg CpG ODN(联合佐剂组)滴鼻,间隔2周,免疫2次。末次免疫后第20 d颈椎脱臼处死小鼠,收集血清和肠冲洗液,ELISA法检测血清弓形虫特异性IgG和肠冲洗液IgA。分离脾淋巴细胞,用四甲基偶氮唑盐(MTT)法检测致敏脾淋巴细胞体外增殖活性。结果ELISA法测定PBS组、抗原组、CpG佐剂组、CT佐剂组及联合佐剂组小鼠血清IgGA值分别为0.025 6±0.012 1、0.054 2±0.024 4、0.109 4±0.007 5、0.110 1±0.009 1和0.124 6±0.013 7,肠冲洗液IgAA值分别为0.141 8±0.030 2、0.415 2±0.080 70、.526 9±0.075 40、.846 5±0.115 5和1.128 9±0.100 3,脾淋巴细胞刺激指数SI分别为1.159 0±0.275 5、1.364 0±0.349 8、1.582 0±0.375 3、1.928 0±0.278 9和2.371 0±0.646 9,差异均有统计学意义(P<0.01);联合佐剂组肠冲洗液IgA水平和脾淋巴细胞的SI值与CpG佐剂组比较差异有统计学意义(P<0.05)。结论STAg联合CpG ODN和/或CT鼻内免疫小鼠可有效诱导机体免疫反应,两种佐剂联合免疫的效果优于单一抗原或抗原联合单一佐剂。

Objective To investigate the adjuvant effect of CpG oligodeoxynucleoties（CpG ODN） and cholera toxin（CT） by observing the IgG in the sera,IgA in intestinal washes,and splenic lymphocyte proliferative response of BALB/c mice in vitro following intranasal immunization with STAg plus CpG ODN and/or CT.Methods Sixty 6-week-old BALB/c mice were randomly divided into five groups,which were immunized intranasally two times at 14-day intervals with 20 μl PBS（PBS group）,20 μg STAg（antigen group）,20 μg STAg plus 10 μg CpG ODN（CpG group）,20 μg STAg plus 1 μg CT（CT group） and 20 μg STAg plus 10 μg CpG ODN and 1 μg CT（combined adjuvant group）,respectively.All animals were dislocated from cervical vertebra on the 20th day after the last immunization.The content of IgG antibody in the sera and IgA antibody in intestinal washes were detected by ELISA and the proliferative reaction of splenic lymphocyte in vitro were tested by MTT.Results The level of IgG in the sera of PBS group,antigen group,CpG group,CT group and combined adjuvant group were 0.025 6±0.012 1,0.054 2±0.024 4,0.109 4±0.007 5,0.110 1±0.009 1 and 0.124 6±0.013 7;The level of IgA in intestinal washes 0.141 8±0.030 2,0.415 2±0.080 7,0.526 9±0.075 4,0.846 5±0.115 5 and 1.128 9±0.100 3;The splenic lymphocyte proliferative activity（SI） were 1.159 0±0.275 5,1.364 0±0.349 8,1.582 0±0.375 3,1.928 0±0.278 9 and 2.371 0±0.646 9,respectively.The IgA level in intestinal washes and splenic lymphocyte proliferate activity（SI） in combined adjuvant group were significantly higher than those in antigen group and CpG group（P〈0.01）.Conclusion The immune response of mice immunized intranasally with STAg plus CpG ODN and/or CT can be elicited effectively.Mice immunized with combined adjuvant had much stronger immune response than those with single STAg or STAg plus CpG ODN or CT.