An improved coating for the isolation and quantitation of interferon-gamma in spiked plasma using surface plasmon resonance (SPR).
An improved coating for the isolation and quantitation of interferon-gamma in spiked plasma using surface plasmon resonance (SPR).
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DOI：
10.1016/j.bios.2004.11.008
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发表时间：
2005-09
影响因子：
12.6
通讯作者：
E. Stigter;G. D. de Jong;W. P. van Bennekom
中科院分区：
文献类型：
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作者：
E. Stigter;G. D. de Jong;W. P. van Bennekom
研究了利用表面等离子体共振技术检测血浆中高等电点模型蛋白重组人干扰素-γ(γ)的方法。最初，一些自组装单分子膜(SAM)和水凝胶衍生的SAM涂层被表征为等离子体组分的吸附和解吸性能。接下来，将抗干扰素-γ的单抗MD-2共价连接到性能最好的葡聚糖修饰的硫代十一酸表面。在由羧基修饰的葡聚糖组成的涂层上，观察到当干扰素-γ被注入缓冲液或稀释血浆中时，相互作用行为不同。在缓冲液中注射干扰素-γ的过程中，观察到相互作用过程的加速，并在注射塞子通过后信号继续增加。注射添加干扰素-γ的稀释血浆后，反应增强，但结合过程没有加速。注射完成后，一些结合的物质如预期的那样解吸，导致信号减弱。在不带电荷的右旋糖苷上，抗体修饰的表面与干扰素-γ在血浆和缓冲液中的相互作用相似。在样品注射过程中，根据溶液中存在的干扰素-γ的浓度，反应以结合率增加。当注射完成时，一些结合的物质被从表面洗掉，只有很少的非特定吸附的等离子体成分的贡献是明显的。从测试的涂层来看，未经修饰的葡聚糖包覆的表面等离子体阻抗传感器盘被证明最适合于检测血浆等复杂基质中的干扰素-γ。干扰素-γ在稀释液和缓冲液中的相互作用相当，在缓冲液和100倍稀释液中均可检测到250ngml以上的干扰素-γ。血浆成分的非特异性吸附较低，而特异性干扰素-γ应答相对较高。
A study was initiated to investigate the use of surface plasmon resonance (SPR) for the detection in plasma of a high pI model protein, recombinant human interferon-γ (IFN-γ). Initially a number of self-assembled monolayers (SAMs) and hydrogel-derivatised SAM-coatings were characterised for the adsorptive and desorptive properties of plasma components. Next a monoclonal anti-IFN-γ antibody, MD-2, was covalently attached to dextran-modified mercaptoundecanoic acid surfaces that performed best. On coatings consisting of carboxyl-modified dextran (CMD) a difference in interaction behaviour was observed when IFN-γ was injected in either buffer or diluted plasma. During the injection of IFN-γ in buffer, an acceleration of the interaction process was observed and the signal continued to increase after the injection plug had passed. Upon injection of diluted plasma spiked with IFN-γ, the response increased without acceleration of the binding process. After the injection was finished, some of the bound material desorbed as expected, resulting in a signal decrease. On non-charged dextrans, the interaction between the antibody-modified surface and IFN-γ in either plasma or buffer was similar. During sample injection the response increased with a binding rate depending on the concentration of IFN-γ present in solution. When the injection was finished, some of the bound material was washed away from the surface and only a minor contribution of non-specific adsorbed plasma components was noticeable. From the coatings tested, the non-modified dextran-coated SPR sensor disks prove to be best suited for the detection of IFN-γ in complex matrices like plasma. The interaction of IFN-γ in both diluted plasma and buffer is comparable and concentrations of IFN-γ of 250ngml−1and higher can be detected in both buffer and 100×-diluted plasma. The non-specific adsorption of plasma components is low, whereas the specific IFN-γ response is relatively high.