一种以吖啶酮作为荧光标记物的抗体催化通用检测方法。

A general assay for antibody catalysis using acridone as a fluorescent tag.

作者信息

Reymond J L, Koch T, Schröer J, Tierney E

机构信息

Department of Molecular Biology, Scripps Research Institute, La Jolla, CA 92037, USA.

出版信息

Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4251-6. doi: 10.1073/pnas.93.9.4251.

DOI:10.1073/pnas.93.9.4251

PMID:8633050

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC39521/

Abstract

A simple and highly sensitive catalysis assay is demonstrated based on analyzing reactions with acridonetagged compounds by thin-layer chromatography. As little as 1 pmol of product is readily visualized by its blue fluorescence under UV illumination and identified by its retention factor (Rf). Each assay requires only 10 microliters of solution. The method is reliable, inexpensive, versatile, and immediately applicable in repetitive format for screening catalytic antibody libraries. Three examples are presented: (i) the epoxidation of acridone labeled (S)-citronellol. The pair of stereoisomeric epoxides formed is resolved on the plate, which provides a direct selection method for enantioselective epoxidation catalysts. (ii) Oxidation of acridone-labeled 1-hexanol to 1-hexanal. The activity of horse liver alcohol dehydrogenase is detected. (iii) Indirect product labeling of released aldehyde groups by hydrazone formation with an acridone-labeled hydrazide. Activity of catalytic antibodies for hydrolysis of enol ethers is detected.

摘要

基于通过薄层色谱分析与吖啶酮标记化合物的反应，展示了一种简单且高度灵敏的催化测定法。在紫外光照射下，低至1皮摩尔的产物因其蓝色荧光很容易被观察到，并通过其保留因子（Rf）进行鉴定。每次测定仅需10微升溶液。该方法可靠、廉价、通用，并且可立即以重复形式用于筛选催化抗体文库。给出了三个例子：（i）吖啶酮标记的（S）-香茅醇的环氧化反应。形成的一对立体异构环氧化物在板上得到分离，这为对映选择性环氧化催化剂提供了一种直接筛选方法。（ii）吖啶酮标记的1-己醇氧化为1-己醛。检测到马肝醇脱氢酶的活性。（iii）通过与吖啶酮标记的酰肼形成腙对释放的醛基进行间接产物标记。检测到催化抗体对烯醇醚水解的活性。