REQUIMTE, Departamento de Química, Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa, 2829-516 Caparica, Portugal.
J Mol Recognit. 2010 Sep-Oct;23(5):462-71. doi: 10.1002/jmr.1013.
A novel magnetic support based on gum Arabic (GA) coated iron oxide magnetic nanoparticles (MNP) has been endowed with affinity properties towards immunoglobulin G (IgG) molecules. The success of the in situ triazine ligand synthesis was confirmed by fluorescence assays. Two synthetic ligands previously developed for binding to IgG, named as ligand 22/8 (artificial Protein A) and ligand 8/7 (artificial Protein L) were immobilized on to MNPs coated with GA (MNP_GA). The dimension of the particles core was not affected by the surface functionalization with GA and triazine ligands. The hydrodynamic diameters of the magnetic supports indicate that the coupling of GA leads to the formation of larger agglomerates of particles with about 1 microm, but the introduction of the triazine ligands leads to a decrease on MNPs size. The non-functionalized MNP_GA bound 28 mg IgG/g, two times less than bare MNP (60 mg IgG/g). MNP_GA modified with ligand 22/8 bound 133 mg IgG/g support, twice higher than the value obtained for ligand 8/7 magnetic adsorbents (65 mg/g). Supports modified with ligand 22/8 were selected to study the adsorption and the elution of IgG. The adsorption of human IgG on this support followed a Langmuir behavior with a Q(máx) of 344 mg IgG/g support and K(a) of 1.5 x 10(5) M. The studies on different elution conditions indicated that although the 0.05 M citrate buffer (pH 3) presented good recovery yields (elution 64% of bound protein), there was occurrence of iron leaching at this acidic pH. Therefore, a potential alternative would be to elute bound protein with a 0.05 M glycine-NaOH (pH 11) buffer.
一种新型的基于阿拉伯树胶（GA）包覆的氧化铁磁性纳米粒子（MNP）的磁性载体被赋予了对免疫球蛋白 G（IgG）分子的亲和性。通过荧光分析证实了原位三嗪配体合成的成功。两种先前开发用于结合 IgG 的合成配体，命名为配体 22/8（人工蛋白 A）和配体 8/7（人工蛋白 L），被固定在涂有 GA 的 MNPs 上（MNP_GA）。颗粒核心的尺寸不受 GA 和三嗪配体的表面功能化的影响。磁性载体的水动力学直径表明，GA 的偶联导致颗粒更大的团聚体的形成，约为 1 微米，但三嗪配体的引入导致 MNPs 尺寸减小。未功能化的 MNP_GA 结合了 28mg IgG/g，比裸 MNP（60mg IgG/g）少两倍。用配体 22/8 修饰的 MNP_GA 结合了 133mg IgG/g 载体，比配体 8/7 磁性吸附剂（65mg/g）的值高两倍。选择用配体 22/8 修饰的载体来研究 IgG 的吸附和洗脱。人 IgG 在这种载体上的吸附遵循 Langmuir 行为，Q(máx)为 344mg IgG/g 载体，K(a)为 1.5 x 10(5) M。对不同洗脱条件的研究表明，虽然 0.05 M 柠檬酸盐缓冲液（pH3）表现出良好的回收产率（洗脱结合蛋白的 64%），但在这种酸性 pH 值下会发生铁浸出。因此，一个潜在的替代方案是用 0.05 M 甘氨酸-NaOH（pH11）缓冲液洗脱结合的蛋白质。
