冻融-冻干法制备的流感疫苗脂质体的细胞免疫研究

Cellular immunity of influenza vaccine lyophilized liposome produced by freeze-thawing

分别用冻融-冻干法、薄膜分散-冻干法制备流感疫苗脂质体干粉,通过小鼠肺部免疫后,从细胞免疫水平考察两种方法制备脂质体的免疫原性。将小鼠分为非脂质体疫苗原液组(non-liposome)、薄膜分散-冻干法制备流感脂质体疫苗组、冻融-冻干法制备流感疫苗脂质体组、阳性对照和阴性对照组(n=5)。脂质体疫苗冻干粉组以每只血凝素含量(以H1N1计)6μg肺部免疫,以非脂质体疫苗原液每只血凝素含量6μg腹腔给药作为阳性对照,以未免疫小鼠(PBS给药)为阴性对照。免疫28d,MTT法检测脾淋巴细胞的增殖,流式细胞术检测CD4^+与CD8^+细胞。结果显示:两种方法制备的流感疫苗脂质体冻干粉均可诱导细胞免疫,且免疫效果相当,说明冻融-冻干法有望进一步应用于减毒活疫苗制备。

To investigate the cellular immunogenicity of influenza vaccine liposomes dry powder using the film- dispersion and freeze-thawing. Mice were divided into the non-liposomal influenza vaccine group, film-dispersion prepared liposomal influenza vaccine group, freeze-thawing lyophilized influenza vaccine liposome group, positive control group and negative control group （n = 5）. 6 μg of hemagglutinin of H1N1 subtype per mouse was delivered tracheally to the mice of lyophilized liposomes groups, with the same dose for non-liposomes intraperitoneally delivered groups as the positive control, and PBS intraperotoneal injection group as the negative control. After 28-day of immunization, the proliferationofsplenic lymphocytes was detected by MTT assay; CD4 ＋ cell and CD8 ＋ cell were analyzed by flow cytometry. The dry powder of influenza vaccine liposomes prepared by the above two methods, both induce cellular immunity, with no significant difference in the induced on immunogenicity between the prepared products（ P 〈 0. 05）. The results showed that freeze-thawing method is feasible in the preparation of vaccine liposomes, leading to the attenuated live vaccine liposome preparation.