Institut de biologie intégrative et des systèmes, Pavillon Charles-Eugène-Marchand, Université Laval, Québec, Québec, Canada G1V 0A6.
J Immunol Methods. 2010 Feb 28;353(1-2):148-50. doi: 10.1016/j.jim.2009.12.007. Epub 2009 Dec 24.
Quantification of chemiluminescent signals from a Western blot is routinely used to determine the increase or the decrease in protein expression or modification in cell or tissue extracts. However, although scientists readily agree that such a procedure is not quantitative, it is nonetheless used quantitatively in most publications without appropriate controls that would increase the accuracy of the measurement. Here we reexamined this aspect and found that the primary antibody itself influences the relation between the Western blot signal and the protein amount on the membrane. This relation is non-linear and varies from one antibody to another. In that context, we strongly encourage researchers to use dilution series and calibration curve when quantifying protein by Western blot using chemiluminescent signal.
从蛋白质印迹中化学发光信号的定量分析通常用于确定细胞或组织提取物中蛋白质表达或修饰的增加或减少。然而，尽管科学家们一致认为这样的程序不是定量的，但在大多数出版物中，它仍然被定量使用，而没有适当的对照来提高测量的准确性。在这里，我们重新研究了这一方面，发现一抗本身会影响印迹信号与膜上蛋白质量之间的关系。这种关系是非线性的，并且因抗体而异。在这种情况下，我们强烈鼓励研究人员在使用化学发光信号进行蛋白质印迹定量时，使用稀释系列和校准曲线。
