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Quantifying cell binding kinetics mediated by surface-bound blood type B antigen to immobilized antibodies

机译:定量由表面结合的血液B型抗原介导的固定抗体介导的细胞结合动力学

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摘要

Cell adhesion is crucial to many biological processes, such as inflammatory responses, tumor metastasis and thrombosis formation. Recently a commercial surface plasmon resonance (SPR)-based BIAcore biosensor has been extended to determine cell binding mediated by surface-bound biomolecular interactions. How such cell binding is quantitatively governed by kinetic rates and regulating factors, however, has been poorly understood. Here we developed a novel assay to determine the binding kinetics of surface-bound biomolecular interactions using a commercial BIAcore 3000 biosensor. Human red blood cells (RBCs) presenting blood group B antigen and CM5 chip bearing immobilized anti-B monoclonal antibody (mAb) were used to obtain the time courses of response unit, or sensorgrams, when flowing RBCs over the chip surface. A cellular kinetic model was proposed to correlate the sensorgrams with kinetic rates. Impacts of regulating factors, such as cell concentration, flow duration and rate, antibody-presenting level, as well as pH value and osmotic pressure of suspending medium were tested systematically, which imparted the confidence that the approach can be applied to kinetic measurements of cell adhesion mediated by surface-bound biomolecular interactions. These results provided a new insight into quantifying cell binding using a commercial SPR-based BIAcore biosensor.
机译:细胞粘附对于许多生物过程至关重要,例如炎症反应,肿瘤转移和血栓形成。最近,基于表面等离振子共振(SPR)的商业BIAcore生物传感器已得到扩展,可以确定由表面结合的生物分子相互作用介导的细胞结合。然而,人们对这种细胞结合如何由动力学速率和调节因子定量控制的了解却很少。在这里,我们开发了一种新颖的测定方法,可使用商用BIAcore 3000生物传感器确定表面结合的生物分子相互作用的结合动力学。当RBC在芯片表面流动时,使用具有B型血抗原的人红细胞(RBC)和带有固定化抗B单克隆抗体(mAb)的CM5芯片来获得响应单位或传感图的时程。提出了细胞动力学模型,以使传感图与动力学速率相关。系统地测试了诸如细胞浓度,流动持续时间和速率,抗体呈递水平以及悬浮液的pH值和渗透压等调节因素的影响,这使该方法可用于细胞的动力学测量由表面结合的生物分子相互作用介导的粘附。这些结果为使用基于商业SPR的BIAcore生物传感器定量细胞结合提供了新的见解。

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