乳腺癌前哨淋巴结转移快速诊断的初步研究  

Preliminary study on rapid diagnosis of sentinel lymph node metastases in breast cancer

作　　者：郝晓鹏[1] 刘毅[1] 孙恒[1] 胡海旭[1] 刘兵[1] 尉承泽[1] 

机构地区：[1]中国人民解放军307医院普通外科,北京100071

出　　处：《中华乳腺病杂志（电子版）》2012年第4期13-17,共5页Chinese Journal of Breast Disease(Electronic Edition)

基　　金：北京市科技计划项目(D09050703570905)

摘　　要：目的建立快速准确、全面客观的判断前哨淋巴结转移状态的分子病理检查方法。方法合成细胞角蛋白19(CK19)的环介导等温扩增(LAMP)引物,应用LAMP技术扩增乳腺癌细胞MCF-7的cDNA,验证引物序列的可靠性;通过RT-LAMP技术分别检测MCF-7、Daudi、Namawal细胞的RNA,验证实验体系的可行性;将手术获取的乳腺癌患者前哨淋巴结行RT-LAMP检测CK19表达,与常规病理学方法的检测结果相对比,最终通过荧光定量以及离心观测沉淀等手段缩短整体检测时间。结果应用LAMP技术能够检测到MCF-7乳腺癌细胞的CK19表达,而Daudi、Namawal细胞CK19的表达均为阴性;经病理检查证实为癌转移阳性的3枚前哨淋巴结,通过RT-LAMP均检测到CK19表达;经荧光定量及离心观测沉淀等手段优化后反应时间可以缩短至1h之内。结论 LAMP检测CK19表达,能够准确区分上皮来源细胞和淋巴瘤细胞,检测转移阳性淋巴结的结果与常规病理学结果完全符合,其具有简便、准确、快速等优点。Objective To establish a rapid and accurate method to objectively detect the metastasis of sentinel lymph node in breast cancer. Methods The loop-mediated isothermal amplification （LAMP） primers of cytokeratin 19 （CK19） was synthetized, LAMP was applied to amplify breast cancer cell line MCF-7 cDNA, and the reliability of the primer sequences was validated. RT-LAMP was applied to detect the RNA sequence in MCF-7, Daudi, and Namawal cells for the further verification of experiment system. Sentinel lymph nodes from breast cancer patients, which were dissected in surgery, underwent RT-LAMP for detection of CK19 expression, then the results were compared with those in routine pathological tests. The overall detection time can be shortened by fluorescence quantitative method and observing precipitation after centrifugation. Results The CK19 expression in breast cancer MCF-7 cells was detected by LAMP while CK19 expression were negative in Daudi, and Namawal cells. Three positive sentinel lymph node with cancer metastasis confirmed by pathological examination presented CK19 expression by RT-LAMP. The reaction time was shortened to within 1 h by the optimization of fluorescent quantitative method and observing precipitation after centrifugation. Conclusions LAMP for detection of CK19 expression can accurately distinguish lymphoma cells from epithelial cells, and the detection results are fully in line with the routine pathological results. This method is simple, accurate and fast.

关 键 词：环介导的等温扩增 细胞角蛋白19 乳腺肿瘤 前哨淋巴结 转移 

分 类 号：R737.9[医药卫生—肿瘤]