Díez Montoro R, Salabert Salvador M T, Moreno Frigols J L
Department of Physical Chemistry, Faculty of Pharmacy, Valencia, Spain.
J Pharm Biomed Anal. 2008 Jun 9;47(2):435-41. doi: 10.1016/j.jpba.2008.01.023. Epub 2008 Jan 19.
Competitive protein binding radioimmunoassay (CPB-RIA) is a principal method for quantifying serum digoxin concentration. The accuracy of this method is critically dependent on factors that influence the substitution reaction between unlabelled (Q) antigen (digoxin) with (125)I-labelled antigen (M) bound to anti-digoxin antibody (P). We studied the influence of initial concentration of M, ionic strength, and viscosity on the substitution reaction between M and Q. In addition, we propose a kinetic model for this reaction.
We used a commercially available CPB-RIA for digoxin, a gamma counter, and a viscosimeter to study the effect of initial concentration of M, ionic strength, viscosity, and temperature on the substitution reaction between M and Q. Data were analyzed using Statistica software.
The apparent rate constant for the reaction between M and Q in the formation of PM is dependent on the initial concentration of M, and the ionic strength, viscosity, and temperature of the reaction medium, and independent of the concentration of Q.
A kinetic model for the displacement of the (125)I-digoxin by the digoxin in its union to a specific antibody is proposed. Such model adjusts satisfactorily to the results and allows the prediction of the calibration curves of RIA (activity bound to the antibody vs. concentration of digoxin) showing the influence of the concentration of both species, the time of incubation, the viscosity and the ionic strength of the medium, on the sensitivity of the method of RIA on which the analytical determination of the digoxin is based.
竞争性蛋白结合放射免疫测定法（CPB - RIA）是定量测定血清地高辛浓度的主要方法。该方法的准确性关键取决于影响未标记（Q）抗原（地高辛）与结合于抗地高辛抗体（P）上的（125）I标记抗原（M）之间置换反应的因素。我们研究了M的初始浓度、离子强度和粘度对M与Q之间置换反应的影响。此外，我们提出了该反应的动力学模型。
我们使用市售的地高辛CPB - RIA、γ计数器和粘度计，研究M的初始浓度、离子强度、粘度和温度对M与Q之间置换反应的影响。使用Statistica软件分析数据。
在形成PM的过程中，M与Q反应的表观速率常数取决于M的初始浓度、反应介质的离子强度、粘度和温度，且与Q的浓度无关。
提出了地高辛在与特异性抗体结合时置换（125）I - 地高辛的动力学模型。该模型能很好地拟合结果，并可预测放射免疫测定法（与抗体结合的活性对地高辛浓度）的校准曲线，显示出两种物质的浓度、孵育时间、介质的粘度和离子强度对作为地高辛分析测定基础的放射免疫测定法灵敏度的影响。
