免疫质谱法检测肝癌血清多肽标志物  

Determination of Hepatocellular Carcinoma Serum Polypeptide Based on Antibody Preconcentration Coupled with Mass Spectrometry

作　　者：傅海媛[1,2] 原剑[2,3] 周晓明[2,3] 孙云波[2,3] 黄亚娟[2,3] 侯利平[2,3] 宋纯艳[2,3] 杨保安 甄蓓[2,3] 高蓉[5] 貌盼勇[5] 林利忠 徐淑芳 肖汉族 魏开华[2,3] 

机构地区：[1]解放军301医院,北京100853 [2]军事医学科学院放射与辐射医学研究所,北京100850 [3]北京蛋白质组研究中心 蛋白质组学国家重点实验室,北京102206 [4]北京正旦国际科技有限责任公司,北京102206 [5]解放军302医院,北京100039 [6]湖南金健药业有限责任公司,常德415001

出　　处：《分析化学》2012年第12期1803-1806,共4页Chinese Journal of Analytical Chemistry

基　　金：卫生部传染病重大专项(No.2008ZX10002-016);863项目(No.2012AA020205)资助

摘　　要：建立免疫亲和富集分离与质谱分析相结合的免疫质谱法用于血清多肽标志物pep5的检测。多克隆抗体与蛋白A琼脂糖颗粒偶联用于捕获pep5,基质辅助激光解析电离飞行时间质谱检测标志物。本方法的线性范围为1～48 nmol/L,检出限0.2 nmol/L;回收率为93.0%～103.1%,相对标准偏差(RSD)为6.4%～11.8%;反复冻融3次pep5,检测结果稳定;检测临床肝癌/正常血清样本灵敏度为78.0%、特异度为90%。该方法方便、快速、准确,适用于临床样本中低浓度标志物的检测研究,对于癌症的早期诊断具有重要意义。Polypeptide antibody pre-concentration method coupled with mass spectrometry detecting was established for the determination of serum polypeptide.Polypeptide was captured by agarose beads followed by binding of specific polyclonal antibodies from serum.After timed washing steps,the remaining bound polypeptides were eluted from the beads and detected by matrix-assisted laser desorption/ionization time-of-flight（MALDI-TOF） mass spectrometry（MS）.We optimized a protein A agarose bead-based platform amenable to high throughput peptide capture followed by mass spectrometry can achieve ion signal,with precision（RSD12%） and accuracy（about 10% of relative error） sufficient for quantifying biomarkers in the physiologically relevant nmol/L range.The method for the detection of peptide 5（pep5） in the clinical hepatocellular carcinoma serum had sensitivity of 78.0% and specificity of 90%.The method is appropriate for the detection of low concentration biomarkers in clinical samples,and it will be significant for earlier diagnosis of cancers.

关 键 词：多肽抗体 免疫亲和富集 质谱 

分 类 号：R735.7[医药卫生—肿瘤] O657.63[医药卫生—临床医学]