Department of Chemistry and Chemical Biology, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA.
Anal Bioanal Chem. 2011 Jan;399(2):773-82. doi: 10.1007/s00216-010-4170-1. Epub 2010 Sep 9.
The preparation and characterization of heparin-immobilized microspheres which were used to bind acidic fibroblast growth factor (aFGF), vascular endothelial growth factor (VEGF), monocyte chemoattractant protein 1 (MCP-1/CCL2), and regulation upon activation normal T cell express sequence (RANTES/CCL5) is described. These beads were used as trapping agents in microdialysis sampling experiments in a separate study. Both free heparin and a synthesized heparin-albumin conjugate were immobilized onto microspheres and compared for their effectiveness. The heparin-albumin conjugate microspheres exhibited significant nonspecific adsorption which appeared to be due to the albumin content. The prepared heparin-immobilized microspheres were stable for 3 months at 4 °C. A bead-based flow cytometric assay was developed to study the binding capacity and specificity of the heparin-immobilized microspheres to cytokines. These heparin-immobilized microspheres exhibited broad dynamic ranges for binding to the four cytokines (aFGF, 1.0-1,000 ng/mL; VEGF, 0.5-1,000 ng/mL; CCL2, 1.95-1,000 ng/mL; CCL5, 1.95-500 ng/mL). Fast binding kinetics of the cytokines to the heparin-immobilized beads suggests that these beads may be useful as affinity agents in microfluidic flow systems.
本文描述了肝素固定化微球的制备和特性，这些微球用于结合酸性成纤维细胞生长因子（aFGF）、血管内皮生长因子（VEGF）、单核细胞趋化蛋白 1（MCP-1/CCL2）和调节激活正常 T 细胞表达序列（RANTES/CCL5）。这些珠子在另一项微透析采样实验中被用作捕获剂。游离肝素和合成的肝素-白蛋白缀合物都被固定到微球上，并比较了它们的效果。肝素-白蛋白缀合物微球表现出明显的非特异性吸附，这似乎是由于白蛋白含量所致。制备的肝素固定化微球在 4°C 下稳定 3 个月。开发了一种基于珠子的流式细胞术测定法来研究肝素固定化微球与细胞因子的结合能力和特异性。这些肝素固定化微球对四种细胞因子（aFGF，1.0-1,000ng/mL；VEGF，0.5-1,000ng/mL；CCL2，1.95-1,000ng/mL；CCL5，1.95-500ng/mL）具有较宽的动态结合范围。细胞因子与肝素固定化珠的快速结合动力学表明，这些珠子可能在微流控流动系统中作为亲和剂有用。
