中文摘要：

目的：通过对已构建的肺腺癌相关人源单链抗体库进行筛选,得到抗过氧化物酶PeroxiredoxinⅠ（PrxⅠ）肺腺癌单链抗体,评价抗体在动物模型体内分布及靶向显像作用。方法：PCR法检测大肠杆菌TG1中单链抗体single-chain variable fragment（scFv）基因插入率;凝胶电泳鉴定SfiⅠ和NotⅠ双酶切质粒结果;以肺腺癌细胞株A549和在肺癌中高表达的抗氧化蛋白PrxⅠ为靶抗原,对抗体库进行富集筛选。阳性克隆用IPTG诱导表达并检测。放射性核素131I标记纯化抗体,进行荷瘤裸鼠体内分布及SPECT放射免疫显像研究。结果：scFv基因插入率为77%（23/30）,双酶切鉴定证实目的条带。通过亲和筛选,肺腺癌单链抗体得到富集,收获率逐轮提高,第6轮是第1轮的180倍。选取10个克隆经ELISA法检测,其中6个克隆与A549细胞呈阳性反应,阳性率60%（6/10）。SDS-PAGE及ELISA检测证实得到人源抗PrxⅠ肺腺癌单链抗体。竞争ELISA显示抗体能有效结合A549细胞。核素标记抗体的放射化学纯度为（95.6±3.7）%,比活度为（2.8±0.2）TBq·g-1。体内分布研究显示抗体与肺腺癌组织有效结合。尾静脉注射48h后,瘤/血和瘤/肌肉放射性比值均达最大值,分别为（4.06±0.13）和（5.17±0.97）%ID·g-1。SPECT示抗体与肿瘤有效结合。抗体注射后48h的T/NT值（3.73±0.20）明显高于12h（1.26±0.15）、24h（2.18±0.16）和72h（2.85±0.18）（均P〈0.01）。结论：利用肺腺癌细胞A549和PrxⅠ为靶抗原,从噬菌体抗体库中筛选获得具有较高特异性的抗PrxⅠ肺腺癌单链抗体。在体内抗体能与肺腺癌组织特异性结合。

英文摘要：

Objective To sieve out the fully human,Peroxiredoxin Ⅰ （Prx Ⅰ）-specific single chain variable fragment （scFv）antibodies from the lung adenocarcinoma-related phage antibody library and analysis the biodistribution of the antibodies in vivo.Methods The insert ratio of scFv antibodies library was identified by polymerase chain reaction（PCR）and the products digested by SfiⅠ/NotⅠ were analyzed on 1%agarose gel. Panning against lung adenocarcinoma cell line A549and Prx Ⅰ were performed three rounds,separately.The positive clones were chosen for soluble expression and analyzed by sodium dodecyl sulfate polyacrylamide gel electropheresis（SDS-PAGE）,enzyme-linked immunosorbent assay（ELISA）and competition ELISA.The tumor-beared athymic mice were injected via the tail vein with purified131I-labeled scFv.The biodistribution analysis and radioimmunoimaging were taken to evaluate the specificity and distribution of antibodies invivo.Results The ratio of recombinant bacteria inserted with scFv was 77% （23/30）and the enzyme digest reaction showed the aim products on 1%agarose gel.From the first to the sixth round of panning,the obtained phages number increased 180times.The positive reactions to A549cells were detected in 6of 10clones.The positive ratio was 60% （6/10）. The human scFv vsagainst PrxⅠof lung adenocarcinoma were confirmed by SDS-PAGE and ELISA.Competition ELISA showed significant,concentration-dependent affinity of scFv antibodies to A549cells.The radiochemical purity of 131I-labeled scFv was （95.6±3.7）% and the specific activity was （2.8±0.2）TBq·g-1.The radiolocalization indexes（RI）of tumor/serum and tumor/muscle were gradually increased,reaching its peak（4.06± 0.13and 5.17±0.97,respectively）at 48hpostadministration.The radioactivity was aggregated in tumor locations and the tumor imaging was clearly observed by single photon emission computed tomography（SPECT）imaging. The T/NT value at 48h（3.73±0.20）was significantly higher than those at 12h（1.2