Shimura K, Karger B L
Barnett Institute, Northeastern University, Boston, Massachusetts 02115.
Anal Chem. 1994 Jan 1;66(1):9-15. doi: 10.1021/ac00073a004.
A new highly sensitive microscale analytical procedure called affinity probe capillary electrophoresis (APCE) is presented. One of the two species, which can form a biospecific complex, is labeled with a fluorescent dye. The affinity probe is used to detect the analyte as a complex after the separation of the excess free probe by capillary electrophoresis. As an example of this approach, an FAB' fragment of a mouse monoclonal antibody (anti-human growth hormone) was labeled with tetramethylrhodamine-iodoacetamide at a hinge region thiol group. Samples were mixed with the purified labeled antibody fragment, and the associated complex was separated by capillary isoelectric focusing with detection by laser-induced fluorescence. In these preliminary results, methionyl recombinant human growth hormone (met-rhGH) could be successfully determined down to detection levels of approximately 5 x 10(-12) M. Mono- and dideamidated variants of met-rhGH were detected simultaneously with the nondeamidated form of the antigen as separate peaks.
介绍了一种名为亲和探针毛细管电泳（APCE）的新型高灵敏度微尺度分析方法。两种能够形成生物特异性复合物的物质之一，用荧光染料进行标记。通过毛细管电泳分离过量的游离探针后，亲和探针用于检测作为复合物的分析物。作为该方法的一个实例，小鼠单克隆抗体（抗人生长激素）的FAB'片段在铰链区硫醇基团处用四甲基罗丹明碘乙酰胺进行标记。将样品与纯化的标记抗体片段混合，通过毛细管等电聚焦分离相关复合物，并通过激光诱导荧光进行检测。在这些初步结果中，甲硫氨酰重组人生长激素（met-rhGH）能够成功测定至约5×10⁻¹² M的检测水平。met-rhGH的单脱酰胺和双脱酰胺变体与抗原的未脱酰胺形式同时被检测为单独的峰。
