Quantification of fragments of human serum inter-α-trypsin inhibitor heavy chain 4 by a surface-enhanced laser desorption/ionization-based immunoassay
Quantification of fragments of human serum inter-α-trypsin inhibitor heavy chain 4 by a surface-enhanced laser desorption/ionization-based immunoassay
复制标题
DOI：
10.1373/clinchem.2005.065722
复制
发表时间：
2006-06-01
影响因子：
9.3
通讯作者：
Zhang, Zhen
中科院分区：
文献类型：
--
作者：
Song, Jin;Patel, Manisha;Zhang, Zhen
背景：通过表面增强或基质辅助激光解吸/电离飞行时间质谱（SELDI-TOF-MS 或 MALDI-TOF-MS），已将人间 α-胰蛋白酶抑制剂重链 4 (ITIH4) 富含脯氨酸区 (PRR) 的几个蛋白水解衍生片段鉴定为潜在的疾病标志物。 方法：此前，我们开发了一种基于 SELDI 的免疫测定法，可以同时区分和量化感兴趣的蛋白质/肽的多个亚型/变体。在本研究中，我们使用这种高通量方法来量化和表征人血清 ITIH4 PRR 内的广泛碎片，并确定其与不同疾病状况的关联。 ITIH4 相关片段首先通过使用与肽特异性抗体偶联的珠子进行免疫捕获。然后通过 SELDI-TOF-MS 对洗脱液进行研究。此外，还使用新鲜收集并立即处理的血清和血浆样本来分析这些ITIH4片段的离体稳定性。结果：人血清ITIH4被证明在PRR内进行了广泛的蛋白水解加工，其片段模式与不同的疾病状况密切相关。断裂模式通常与内切蛋白酶裂解和外切蛋白酶作用一致。观察到的碎片在不同的检测条件或血液采集和处理程序下变化不大。结论：人血清ITIH4 PRR内的碎片模式与不同的疾病状况相关，并且可能包含重要的诊断信息。这些碎片模式可用作癌症检测和分类的潜在生物标志物。 (c) 2006 年美国临床化学协会。
Background: Several proteolytically derived fragments from the proline-rich region (PRR) of human inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4) have been identified by surf ace-enhanced or matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS or MALDI-TOF-MS) as potential disease markers.Methods: Previously, we developed a SELDI-based immunoassay that can simultaneously distinguish and quantify multiple isoforms/variants of a protein/peptide of interest. In this study, we used this high-throughput approach to quantify and characterize the extensive fragmentation within the PRR of human serum ITIH4 and determined its association with different disease conditions. The ITIH4-related fragments were first immunocaptured by use of beads coupled with peptide-specific antibodies. The eluates were then studied by SELDI-TOF-MS. In addition, freshly collected and immediately processed serum and plasma samples were used to analyze the ex vivo stability of these ITIH4 fragments.Results: Human serum ITIH4 was shown to be extensively proteolytically processed within the PRR, and its fragmentation patterns were closely associated with different disease conditions. Fragmentation patterns were generally consistent with cleavages by endoprotease followed by exoprotease actions. Observed fragments changed little under different assay conditions or blood collection and processing procedures.Conclusions: The fragmentation patterns within the PRR of human serum ITIH4 are associated with different disease conditions and may hold important diagnostic information. These fragmentation patterns could be useful as potential biomarkers for detection and classification of cancer. (c) 2006 American Association for Clinical Chemistry.