摘要
目的观察抗小鼠TLR-2胞外段B细胞识别表位抗体对小鼠肉瘤S180生长的影响。方法在对小鼠Toll样受体2(mTLR-2)B细胞优势表位预测的基础上,合成B细胞识别表位20mer短肽,将其与载体蛋白偶联,制备免疫原。所得兔抗mTLR-2胞外段B细胞识别表位多克隆抗体TSP-1纯化后,采用Westernblotting、免疫组织化学和流式细胞仪进行初步鉴定。小鼠肉瘤株S180细胞注射小鼠左后肢,其中实验组混合100μg纯化抗体TSP-1,无关免疫球蛋白对照组混合100μg正常兔IgG,空白对照组仅接种S180。小鼠经麻醉分别于10、14、18d处死,称量瘤质量并计算抑瘤率。结果Westernblotting显示在相对分子质量约95000处可见清晰的反应条带;免疫组织化学和流式细胞术检测显示抗体可与表达天然mTLR-2的J774A.1细胞反应;实验组小鼠肉瘤生长明显受到抑制,其瘤质量在10、14、18d与对照组相比差异显著(P<0.05);抑瘤率分别为77%、51%和53%。结论局部应用抗mTLR-2胞外段B细胞识别表位抗体可抑制小鼠肉瘤S180生长,且表现在肿瘤生长的早期。
Objective To observe the effect of the antibody to a B cell epitope on mouse Toll-like receptor-2 (mTLR-2) extracellular domain on the growth of murine fibrosarcoma. Methods An immunogen was prepared by conjugating a 20-mer peptide, synthesized on the basis of prediction for B cell dominant epitope of mTLR-2, to a carrier protein. Rabbit polyclonal antibodies against B cell epitope of mTLR-2 were prepared and purified, and characterized by Western blotting, immunohisto- chemistry and flow cytometry. Murine fibrosarcoma S180 cells were inoculated into the left hindlimbs of Swiss mice, and in the treatment group, the injected cells were mixed with 100 γg anti-mTLR-2 antibody TSP-1, with the same dose of irrelevant rabbit IgG in the IgG control group and with buffer solution only in the blank control group. The mice in each group were anesthetized and sacrificed on days 10, 14, and 18 following the injections, respectively, and the weight of the tumors was measured and the inhibition rate calculated. Results A distinct band for the antibody TSP-1 was shown at the relative molecular mass of 95 000 by Western blotting. The antibody TSP-1 could also react with native mTLR-2 expressed on J774A.1 cells, as identified by immunohistochemistry and flow cytometry. The growth of the fibrosarcoma S180 was obviously inhibited by injections with the antibody TSP-1 mixture, and the weight of tumor in mice treated with TSP-1 was significantly lower than that in the two control groups (P<0.05), with an inhibition rate of 77% on day 10, 51% on day 14 and 53% on day 18. Conclusion Local application of the antibody against B cell epitope on mTLR-2 extracellular domain can inhibit the growth of murine fibrosarcoma, especially in the early stage of tumor proliferation.
出处
《第一军医大学学报》
CSCD
北大核心
2004年第8期873-876,共4页
Journal of First Military Medical University
基金
国家自然科学基金(30070719)~~
