DNA免疫诱导小鼠产生抗人心肌肌钙蛋白T抗血清的研究

目的 通过基因免疫诱导BALB／c小鼠产生抗人心肌肌钙蛋白T(cTnT)的抗体应答，并比较不同免疫佐剂、预处理方法对免疫应答的影响。方法 构建cTnT的真核表达质粒pCI-cTnT，大量提取、PEG纯化质粒，通过肌肉注射免疫BALB／c小鼠并加强免疫，每隔2周取血，ELISA法分析抗cTnT的体液应答，免疫印迹检测抗血清的特异性。并比较布吡卡因预处理、CpG ODN核酸佐剂、弗氏不完全佐剂对抗体应答效果的影响。结果 接种pCI-cTnT质粒的小鼠血清中检测出抗cTnT特异性抗体，加强免疫使抗体滴度提高；使用CpGODN后免疫应答增强，弗氏不完全佐剂可明显增强免疫效果，而吡卡因预处理使应答水平下降；初次免疫后14周仍可测得较高的抗体应答。结论 采用含cTnT基因的质粒DNA免疫，成功地诱导小鼠产生抗cTnT的抗体；传统的弗氏不完全佐剂可作为核酸免疫的佐剂使用。

DNA immunization with recombinant expression plasmid of human cardiac troponin T

Objective To induce an antibody response to human cardiac troponin T (cTnT) by intramuscular immunization with recombined expression plasmid DNA for cTnT and to compare the efficacy of different adjuvants. Methods cDNA of cTnT gene was cloned into mammalian expression vector and the expression plasmids were purified by PEG method. Specific antibodies in sera were analyzed by ELISA and Western blot after the cTnT expression plasmids were intramuscularly injected into mice. The efficacy of different adjuvant/pretreatment methods: CpG-ODN, pretreatment with bupivacaine, incomplete Freund's adjuvant (IFA), on the immunogenicity of DNA vaccines were also assessed. Results Specific antibody to cTnT was induced by DNA-mediated immunization. The immunogenicity of pCI-cTnT vector was increased by using CpG-ODN while reduced by bupivacaine pretreatment method. IFA has a significant effect on antibody responses of gene immunization. Conclusion Specific antibody response to human cTnT was successfully induced by DNA immunization IFA can be used as an adjuvant in DNA immunization.