细胞表位的研究.ppt

* * THANK YOU SUCCESS * * 可编辑 1. SARS-CoV S DNA 疫苗 Zhi-yong Yang (NIH) Nature. 2004, 428 (1 ) 2.免疫方法 8w female BALB/c prime at week 0 50ug DNA boost twice at week 3, 6 2-3w after prime 4-6w after boost 1-2w after boost to observe the effect of prime immunization to observe the effect of boost immunization to observe the effect of long term immunization :mouse sacrificed 50ug DNA i.m i.m 3. 单细胞悬液的准备 淋巴结 脾 肺 剪碎（消化），研磨，200目筛网过滤， 裂解红细胞，2×106/ ml 4. SARS-CoV S蛋白多肽 169条, 17-19aa, 10aa重叠 (NIH NIAID VRC) P1 S1-17 P2 S8-25 P3 S16-22 P168 S1200-1216 P169 S1207-1255 1. S抗原刺激以后IFN-? （图A）和IL-2的产生（图B） 结 果 与 讨 论 The empty circles represent the results in the absence of peptides and solid circles represent the results in the presence of peptides. * * Fig. 2. Verification of potential SARS CoV S epitopes Potential SARS CoV S epitopes P50, P51, P59 and P60 in pool 3, and P141, P144, P151 and P152 in pool 8 were used to stimulate splenocytes from SARS CoV S DNA immunized BALB/c mice. IFN-γ production was detected by ELISA (A) and ELISPOT (B), respectively. Each symbol represents the results of an individual experiment (n=3~7). In addition, intracellular cytokine staining (C) was performed to determine CD4+ or CD8+ T cell population. “0” represents the non-peptide stimulated control. Numbers at the corner in each sample represent the percentage of positive cells. Representative results of three independent experiments were shown. * Fig. 3. Identification of new synthetic 10aa CD4 and CD8 epitopes The overlapped amino acids between P50 and P51 (N50), and between P59 and P60 (N60) were synthesized. Peptides N50 and N60 were used to stimulate splenocytes from SARS CoV S DNA vaccine-immunized mice. P50 and P60 were used as positive controls, respectively. ELISA (A) and ELISPOT (B) were performed as described above. Furthermore, N50 and 60 were serial diluted to stimulate splenocytes from the DNA immunized mice, ELISPOT (C) was performed to detect numbers o