Keener C R, Wolfe C A, Hage D S
University of Nebraska, Lincoln.
Biotechniques. 1994 May;16(5):894-7.
The oxidation of antibody carbohydrate residues is a common approach used for the site-specific immobilization or modification of antibodies. One way of following this oxidation process is to label the resulting aldehyde groups with a dye such as Lucifer yellow CH (LyCH). This study examined the optimum conditions for preparing and purifying antibody-LyCH conjugates. A 250-fold excess of LyCH reacted with antibody at pH 6.5 for two or more hours gave maximum labeling. Nonreacted LyCH could be effectively removed by passing the labeled antibody through a size exclusion column, followed by one or two dialysis cycles. The LyCH antibody conjugates were found to be stable for at least three weeks when stored in pH 7.4 phosphate buffer.
抗体碳水化合物残基的氧化是用于抗体位点特异性固定或修饰的常用方法。追踪这种氧化过程的一种方法是用诸如路西法黄CH（LyCH）之类的染料标记生成的醛基。本研究考察了制备和纯化抗体-LyCH缀合物的最佳条件。在pH 6.5下，250倍过量的LyCH与抗体反应两小时或更长时间可实现最大标记。通过将标记的抗体通过尺寸排阻柱，然后进行一到两个透析循环，可以有效去除未反应的LyCH。发现LyCH抗体缀合物在pH 7.4磷酸盐缓冲液中储存时至少可稳定三周。
