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Assay of chloramphenicol acetyl transferase by high-performance liquid chromatography
L O Burzio1, M Brito, A M Zarraga
1Instituto de Bioquimica, Facultad de Ciencias, Universidad Austral de Chile, Valdivia.
Abstract:
A procedure to measure chloramphenicol acetyl transferase (CAT) activity by reverse-phase high-performance liquid chromatography is described. The antibiotic as well as the acetylated derivatives are well resolved on a Superspher RP-18 column using equal parts of acetonitrile and 10 mM sodium acetate (ph 5.0) as a solvent. Under these conditions, less than 100 pmol of each derivative can be easily detected within 10 minutes, and no radioactive chloramphenicol is needed. The present procedure has been used to measure the activity of the enzyme in extracts of chicken fibroblast transfected with the recombinant plasmid pSV2-cat containing the CAT gene.