Analysis of native proteins from biological fluids by biomolecular interaction analysis mass spectrometry (BIA/MS): exploring the limit of detection, identification of non-specific binding and detection of multi-protein complexes.
Analysis of native proteins from biological fluids by biomolecular interaction analysis mass spectrometry (BIA/MS): exploring the limit of detection, identification of non-specific binding and detection of multi-protein complexes.
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DOI：
10.1016/s0956-5663(01)00229-9
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发表时间：
2001-12
影响因子：
12.6
通讯作者：
D. Nedelkov;Randall W. Nelson
中科院分区：
文献类型：
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作者：
D. Nedelkov;Randall W. Nelson
生物分子相互作用分析质谱（BIA/MS）是一种二维分析技术，可以定量和定性地检测感兴趣的分析物。在第一个维度，表面等离子体共振（SPR）被用于检测生物分子在其天然环境。由于SPR检测是非破坏性的，因此保留在SPR主动传感器表面的分析物可以使用基质辅助激光解吸/电离飞行时间（MALDI-TOF）质谱法在第二次元中进行分析。MALDI-TOF MS分析的定性性质补充了SPR检测的定量特征，克服了SPR检测无法区分和表征多蛋白复合物和非特异性结合的缺点。在这项工作中，建立了SPR传感后进行MS分析的好处。研究人员对生物体液中存在的四种标志物（胱抑素C、β -2微球蛋白、尿蛋白1和视黄醇结合蛋白）的检索和检测进行了探索，以证明BIA/MS在同时检测临床相关生物标志物和描述非特异性结合方面的有效性。此外，还研究了在极低SPR响应下BIA/MS的检出限。最后，利用BIA/MS首次实现了体内组装蛋白复合物的检测。
Biomolecular interaction analysis mass spectrometry (BIA/MS) is a two-dimensional analytical technique that quantitatively and qualitatively detects analytes of interests. In the first dimension, surface plasmon resonance (SPR) is utilized for detection of biomolecules in their native environment. Because SPR detection is non-destructive, analyte(s) retained on the SPR-active sensor surface can be analyzed in a second dimension using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. The qualitative nature of the MALDI-TOF MS analysis complements the quantitative character of SPR sensing and overcomes the shortcomings of the SPR detection stemming from the inability to differentiate and characterize multi-protein complexes and non-specific binding. In this work, the benefit of performing MS analysis following SPR sensing is established. Retrieval and detection of four markers present in biological fluids (cystatin C, beta-2-microglobulin, urinary protein 1 and retinol binding protein) was explored to demonstrate the effectiveness of BIA/MS in simultaneous detection of clinically related biomarkers and delineation of non-specific binding. Furthermore, the BIA/MS limit of detection at very low SPR responses was investigated. Finally, detection of in-vivo assembled protein complexes was achieved for the first time using BIA/MS.