N-甲基-D-天门冬氨酸受体2B亚基抗原表位DNA疫苗免疫大鼠的效应

目的 采用N-甲基-D-天门冬氨酸受体2B亚基抗原表位(eNg2B)插人质粒载体pDC515中制备的DNA疫苗(pDC515/eNR2B)免疫大鼠,评价其免疫效应.方法 成年雄性SD大鼠40只,体重200～250 g,随机分为4组:对照组(C组)、PBS组、pDC515组和pDC515/eNR2B组,每组10只.PBS组、pDC515组和pDC515/eNR2B组分别肌肉注射PBS 100 μl、pDC515 100 μg和pDC515/eNR2B 100 μg进行免疫,1周后以等剂量再次免疫.于第1次免疫前、第2次免疫后8、14、21 d时,截尾法采集尾静脉血,分离血清,ELISA法检测NR2B抗体水平.于第2次免疫后21 d时处死大鼠取脊髓,分别采用免疫组织化学法和Western blot法检测NB2B抗体与自身腰段脊髓组织NR2B特异性结合情况.结果 第2次免疫后8 d,pDC515/eNR2B组血清中可检测到NR2B抗体,14 d时达高峰,并高滴度持续至21 d(P＜0.05);C组、PBS组和pDC515组均未检测到相应NR2B抗体;免疫组织化学法和Western blot法检测到pDC515/eNR2B组血清NR2B抗体可与自身脊髓组织NR2B结合.结论 pDC515/eNR2B可诱导SD大鼠产生体液免疫反应,所产生的自身抗体在血液中可高滴度存在一定时间.

Response of the rats to immunization with DNA vaccine contazining N-methyl-D-aspartate receptor subunit NR2B epitope

Objective To investigate the response of the rats to immunization with DNA vaccine(pDC515/eNR2B)developed by inserting NMDA receptor subunit NR2B epitope(eNB2B)into plasmid vector (pDC.515).Methods Forty adult male SD rats weiighing 200-250 g were randomly divided into 4 groups(n=10 each):group Ⅰ control;groupⅡ PBS;group Ⅲ pDC515 and group Ⅳ pDC515/eNR2B.PBS 100 μl or pDC515 100 μg or pDC515/eNR2B 100 μg was injected into the quadriceps muscle of the rats in group Ⅱ, Ⅲ and Ⅳ respectively once a week for two weeks.Blood samples were obtained for detection of the specific antibody against NR2B in serum before immunization and at day 8,14,21 after second immunization(by ELISA).The ability of the specific antibody against NR2B to combine with its own lumbar spinal cord was determined by immunohistochemistry and Western blot.Results The specific antibody against NR2B was detected in the serum at 8 d after the second immunization in group Ⅳ(pDC515/eNR2B),and reached the peak at 14 d.The high titer lasted until the 21 d.No specific antibody against NR2B was detected in the seram in group Ⅱ and Ⅲ.The specific antibody against NR2B in the serum of the rats immunized with pDCS15/eNR2B could combine with its own lumbar spinal cord indicating that it was an autoantibody.Conclusion The pDC515/eNR2B Can induce humoral immune response in SD rats.The hish titer of the autoantibody produced in the blood can last for a certain period of time.