Karlsson R, Michaelsson A, Mattsson L
Pharmacia Biosensor AB, Uppsala, Sweden.
J Immunol Methods. 1991 Dec 15;145(1-2):229-40. doi: 10.1016/0022-1759(91)90331-9.
An automated biosensor system for measuring molecular interactions has been used to study the kinetics of monoclonal antibody-antigen reactions. The system combines a microfluidic unit in contact with a sensor surface for surface plasmon resonance detection. The specificity of the surface is determined by the operator. Antibody or antigen is immobilised in a dextran matrix attached to the sensor surface. The interaction of matrix bound antibody or antigen with the corresponding partner in solution is monitored in real time. None of the interacting molecules needs to be labelled and it is not necessary to determine the concentration of the the matrix bound component in advance. Two systems were studied: matrix bound monoclonal antibodies (MAbs) interacting with HIV-1 core protein p24 and immobilised aminotheophylline reacting with MAbs. Control of the amount of immobilised ligand and reusable sensor surfaces permits the comparison of different MAbs reacting with antigen under almost identical conditions. Differences in affinity and reaction rates are immediately apparent. The calculated association rate constants for p24 MAbs ranged from 3 x 10(4) - 7.4 x 10(5) M-1 s-1 and for theophylline MAbs association rate constants as high as 1 x 10(6) M-1 s-1 were encountered. The calculated dissociation rate constants were in the region 2 x 10(-4) s-1 to 2 x 10(-2) s-1.
一种用于测量分子相互作用的自动化生物传感器系统已被用于研究单克隆抗体 - 抗原反应的动力学。该系统结合了一个与传感器表面接触的微流体单元，用于表面等离子体共振检测。表面的特异性由操作人员确定。抗体或抗原固定在附着于传感器表面的葡聚糖基质中。实时监测基质结合的抗体或抗原与溶液中相应配体的相互作用。相互作用的分子均无需标记，也无需预先确定基质结合成分的浓度。研究了两个系统：基质结合的单克隆抗体（MAb）与HIV - 1核心蛋白p24相互作用，以及固定化的氨茶碱与单克隆抗体反应。对固定化配体的量和可重复使用的传感器表面的控制使得能够在几乎相同的条件下比较不同的单克隆抗体与抗原的反应。亲和力和反应速率的差异立即显现出来。计算得出的p24单克隆抗体的缔合速率常数范围为3×10⁴ - 7.4×10⁵ M⁻¹ s⁻¹，而茶碱单克隆抗体的缔合速率常数高达1×10⁶ M⁻¹ s⁻¹。计算得出的解离速率常数在2×10⁻⁴ s⁻¹至2×10⁻² s⁻¹范围内。
