Improvement and validation of the fluorescence-based histone deacetylase assay using an internal standard.
The determination of the activity of histone deacetylase (HDAC) and the potency of its inhibitors has become an important goal in medicinal chemistry. This is due both to the involvement of HDAC in gene regulation and the ability of its inhibitors to modulate transcription and induce differentiation and/or apoptosis in cancer cells. We have previously reported the development of a non-isotopic assay for HDAC using a fluorescent derivative of epsilon-acetyl lysine. It can replace existing methods that rely on radioactively labeled histones or oligopeptides as substrates. Here we report validation and improvement of the procedure using an internal standard for the quantitation of the fluorescent substrate by HPLC.
Hoffmann
K
K
Department of Pharmaceutical Chemistry, Westfälische Wilhelms-Universität Münster, Hittorfstr. 58-62, 48149 Münster, Germany.
Heltweg
B
B
Jung
M
M
D000818:Animals; D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D005456:Fluorescent Dyes; D006655:Histone Deacetylases / Q000032:analysis*; D051381:Rats; D012015:Reference Standards; D015203:Reproducibility of Results
D005456:Fluorescent Dyes; D006655:Histone Deacetylases