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. 1996 Apr 30;93(9):4251–4256. doi: 10.1073/pnas.93.9.4251

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

J L Reymond 1, T Koch 1, J Schröer 1, E Tierney 1
PMCID: PMC39521  PMID: 8633050

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.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson J. M. Fluorescent hydrazides for the high-performance liquid chromatographic determination of biological carbonyls. Anal Biochem. 1986 Jan;152(1):146–153. doi: 10.1016/0003-2697(86)90133-8. [DOI] [PubMed] [Google Scholar]
  2. Chayes R., Dvir R., Gould S., Harell A. 1-Dimethylaminonaphthalene-5-sulfonyl hydrazine (dansyl hydrazine): a fluorometric reagent for carbonyl compounds. Anal Biochem. 1971 Jul;42(1):283–286. doi: 10.1016/0003-2697(71)90037-6. [DOI] [PubMed] [Google Scholar]
  3. Fenniri H., Janda K. D., Lerner R. A. Encoded reaction cassette for the highly sensitive detection of the making and breaking of chemical bonds. Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):2278–2282. doi: 10.1073/pnas.92.6.2278. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Kraut J. How do enzymes work? Science. 1988 Oct 28;242(4878):533–540. doi: 10.1126/science.3051385. [DOI] [PubMed] [Google Scholar]
  5. Schultz P. G., Lerner R. A. From molecular diversity to catalysis: lessons from the immune system. Science. 1995 Sep 29;269(5232):1835–1842. doi: 10.1126/science.7569920. [DOI] [PubMed] [Google Scholar]
  6. Seiler N., Wiechmann M. Zur Chromatographie einiger 1-Dimethylamino-naphthalin-5-sulfonyl-derivate auf Kieselgel G-Schichten. J Chromatogr. 1967 Jun;28(2):351–362. doi: 10.1016/s0021-9673(01)85976-8. [DOI] [PubMed] [Google Scholar]
  7. Stewart J. D., Benkovic S. J. Transition-state stabilization as a measure of the efficiency of antibody catalysis. Nature. 1995 Jun 1;375(6530):388–391. doi: 10.1038/375388a0. [DOI] [PubMed] [Google Scholar]
  8. Tawfik D. S., Green B. S., Chap R., Sela M., Eshhar Z. catELISA: a facile general route to catalytic antibodies. Proc Natl Acad Sci U S A. 1993 Jan 15;90(2):373–377. doi: 10.1073/pnas.90.2.373. [DOI] [PMC free article] [PubMed] [Google Scholar]

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