[1]庞华,罗弋,李少林.人源肺腺癌噬菌体抗体库的构建及初步筛选[J].第三军医大学学报,2008,30(22):2082-2085.
 PANG Hua,LUO Yi,LI Shao-lin.Construction of and screening for phage antibody library of human lung adenocarcinoma[J].J Third Mil Med Univ,2008,30(22):2082-2085.
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人源肺腺癌噬菌体抗体库的构建及初步筛选(/HTML )
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
30卷
期数:
2008年第22期
页码:
2082-2085
栏目:
论著
出版日期:
2008-11-30

文章信息/Info

Title:
Construction of and screening for phage antibody library of human lung adenocarcinoma
作者:
庞华 罗弋 李少林
重庆医科大学:附属第一医院核医学科,核医学教研室
Author(s):
PANG Hua LUO Yi LI Shao-lin
Department of Nuclear Medicine, The First Affiliated Hospital, Department of Nuclear Medicine, Chongqing Medical University, Chongqing 400016, China
关键词:
噬菌体抗体库单链抗体肺腺癌筛选
Keywords:
phage antibody library single-chain variable fragment lung adenocarcinoma screening
分类号:
R392.13;R73-35;R734.2
文献标志码:
A
摘要:
目的    构建人源肺腺癌噬菌体单链抗体库,并筛选肺腺癌细胞A549特异性单链抗体。    方法    利用肺腺癌患者癌旁淋巴结组织构建肺腺癌噬菌体单链抗体库。从该抗体库中筛选特异性识别A549细胞的单链抗体,将阳性克隆菌转化E.coli HB2151进行可溶性表达。抗体亲和层析纯化后经SDS-PAGE、Western blot鉴定,通过ELISA法鉴定其与人肺腺癌细胞结合的特异性。    结果    成功构建了1个4.6×107的噬菌体抗体库。在筛选过程中,肺腺癌噬菌体单链抗体得到富集,收获率逐轮得到提高,第5轮为第1轮的181倍。在E.coli HB 2151中实现了单链抗体的可溶性表达。SDS-PAGE与Western blot结果显示抗体相对分子质量为30×103左右。细胞ELISA测定结果显示可溶性抗体具有较高的特异性,能与A549细胞结合,而不与MDA-MB-435细胞结合。    结论    从肺腺癌病人癌肿周围淋巴结扩增免疫球蛋白基因成功地构建人源肺腺癌噬菌体单链抗体库,并从中筛选到肺腺癌特异性抗体。
Abstract:
Objective    To construct human single-chain antibody gene library associated with lung adenocarcinoma, and screen lung adenocarcinoma cell A549-specific antibody.     Methods    The lymphatic tissue near lung adenocarcinoma was used to construct human single-chain antibody gene library. The specific antibody to lung adenocarcinoma cell A549 was screened from the antibody library. Positive clone bacteria were transformed to E. coli HB2151, and their dissolvability was observed. The soluble scFv was purified by affinity chromatography and its relative molecular mass was determined by SDS-PAGE and Western blotting. The specificity of scFv to human lung adenocarcinoma cells was identified with ELISA.     Results    The phage antibody library of 4.6×107 was constructed successfully. Single-chain antibody of human lung adenocarcinoma was enriched. The ratio of yield was increased gradually, 181 times higher after 5 rounds of panning than after the first round of panning. SDS-PAGE and Western blotting results showed soluble scFv’ molecular mass was about 30 000. ELISA showed dissolved antibody had high specificity to bind A549 cells, not MDA-MB-435.     Conclusion    The single-chain antibody gene library of human lung adenocarcinoma was constructed successfully, and specific antibodies to lung adenocarcinoma were screened, providing the basis for single-chain antibody radionuclide imaging and treatment.

参考文献/References:

庞华, 罗弋, 李少林. 人源肺腺癌噬菌体抗体库的构建及初步筛选[J]. 第三军医大学学报,2008,30(22):2082-2085.

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更新日期/Last Update: 2008-10-30