A general assay for antibody catalysis using acridone as a fluorescent tag.
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.
Reymond
J L
JL
Department of Molecular Biology, Scripps Research Institute, La Jolla, CA 92037, USA.
Koch
T
T
Schröer
J
J
Tierney
E
E
D000166:Acridines / Q000138:chemical synthesis / Q000737:chemistry; D000426:Alcohol Dehydrogenase / Q000378:metabolism*; D000818:Animals; D000906:Antibodies; D002384:Catalysis; D002855:Chromatography, Thin Layer / Q000379:methods; D004852:Epoxy Compounds; D005456:Fluorescent Dyes; D006736:Horses; D007202:Indicators and Reagents; D009682:Magnetic Resonance Spectroscopy; D010084:Oxidation-Reduction; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence; D013237:Stereoisomerism
D000166:Acridines; D000906:Antibodies; D004852:Epoxy Compounds; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D000426:Alcohol Dehydrogenase