Multiplexing Determination of Small Cell Lung Cancer Biomarkers and Their Isovariants in Serum by Immunocapture LC-MS/MS
Multiplexing Determination of Small Cell Lung Cancer Biomarkers and Their Isovariants in Serum by Immunocapture LC-MS/MS
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DOI：
10.1021/ac500986t
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发表时间：
2014-07-15
影响因子：
7.4
通讯作者：
Reubsaet, Leon
中科院分区：
文献类型：
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作者：
Torsetnes, Silje B.;Levernaes, Maren S.;Reubsaet, Leon
建立了免疫亲和微球共萃取-选择反应监测（SRM）联合检测小细胞肺癌（SCLC）标志物前胃泌素释放肽（ProGRP）和神经元特异性烯醇化酶（NSE）的方法。的性能进行了比较，最近验证的个人标志物测定与两个IA SRIVI方法。多重方法减少了样品体积、每个样品的处理时间和试剂消耗，并显示出良好的线性、回收率、定量测量，和灵敏度，检测下限（LLOD）值为7.2 pM 5 pM（=210 pg/mL）和定量下限（LLOQ）值分别为24 pM（=300 pg/mL）和15 pM（=700 pg/mL）。该方法的新方面是ProGRP和NSE的多重化，具有通过选择性测定ProGRP同种型1、ProGRP同种型3和总ProGRP以及NSE同工酶的α-和γ-亚基来进行指纹分析的额外能力。分析了来自SCLC患者的六个血清样品，以证明同时区分和单独定量ProGRP、NSE及其同工型和同工酶变体的方法的可行性。所有亚型和同工酶之间的存在和变化，以及与总ProGRP和γ-NSE的常规免疫测定的共变结果，在患者血清样品的分析中观察到。
A multiplex method for the determination of the small cell lung cancer (SCLC) markers progastrin releasing peptide (ProGRP) and neuron specific enolase (NSE) is presented, which involves coextraction by immunoaffinity (IA) beads and codetermination by selected reaction monitoring (SRM). The performance was compared with two IA SRIVI methods which were recently validated for individual marker determination. The multiplexing method reduces sample volume, handling time per sample, and reagent consumption and shows good linearity, recovery, quantitative measurements, and sensitivity with lower limit of detection (LLOD) values of 7.2 pM (=90 pg/rnL) and 4.5 pM (=210 pg/mL) and lower limit of quantitation (LLOQ) values of 24 pM (=300 pg/rnL) and 15 pM (=700 pg/mL), for total ProGRP and gamma-NSE, respectively. The novel aspect of this approach is the multiplexing of ProGRP and NSE with the additional ability to perform fingerprinting by the selective determination of ProGRP isoform 1, ProGRP isoform 3, and total ProGRP, as well as the alpha- and the gamma-subunit of NSE isoenzymes. Six serum samples from patients with SCLC were analyzed to demonstrate the methods feasibility to simultaneously differ between and individually quantify ProGRP, NSE, and their isoform and isoenzyme variants, respectively. Both the presence of and variation between all the isoforms and isoenzymes, as well as covarying results with the conventional immunometric assays for total ProGRP and gamma-NSE, were seen in the analyses of patient serum samples.