Automation of Sample Dispensing using Pocket Tips for Evaluation of CYP2C9 Isoenzyme Reversible Inhibition in an Evolution Precision Pipetting Platform

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• 作者：Manuel Ramí ; rez ; Consuelo Tudela ; Juan B. Crespo ; Alicia Robles ; Antonio Jimé ; nez ; Guillermo Tarazona ; Laura Ferná ; ndez ; Pilar Sá ; nchez ; Doug Kroncke ; Noemí ; Tabanera ; José ; Pé ; rez ; Manuel Sá ; nchez ; José ; R. Tormo ; Fern ; o Pelá ; ez
• 关键词：CYP (cytochrome P-450) isoenzymes ; PocketTips Technology ; PerkinElmer Evolution P3 ; TECAN Ultra Evolution ; TECAN Spectra Fluor ; WinPREP software
• 刊名：Journal of the Association for Laboratory Automation
• 出版年：2008
• 出版时间：October 2008
• 年：2008
• 卷：13
• 期：5
• 页码：289-296
• 全文大小：1233 K

文摘

Most of the medicinal chemistry compounds received from other Merck Research Laboratories (MRL) for evaluation in our in vitro counterscreenings are in pure DMSO solution. This solvent has shown to be a key interference factor in the radiometric evaluation of cytochrome P-450 (CYP) inhibition. To avoid this problem, we have been using a ternary solvent solution that keeps DMSO concentration to a minimum during the titration of the compounds.

The equipment used at our laboratory to perform the dose–response titration of the compounds in the CYP assays is PerkinElmer Evolution Precision Pipetting Platform (EP3) robots with limited microliter capabilities using disposable tips. The need for a nanoliter dispensing technique that keeps the percentage of DMSO at levels that do not affect the assay, and the need to use pure DMSO solutions at high concentrations to avoid solubility issues of the compounds, moved us to design a new automated sample management process for all compounds that were requested to be tested in these assays.

The PocketTips system (nAscent BioSciences Inc., Wellesley Hills, MA) seemed to be a good option for pipetting very low volumes without acquiring new expensive nanoliter liquid handling equipment. Apparent difficulties with implementation were the unavailability of PocketTips for the 96-tips head of the EP3 (PerkinElmer, Waltham, MA) and the need for testing several CYP isoenzymes (up to four) by using the same rack of tips to save run time and reduce costs.

In this article, we discuss the entire process to introduce and validate a new dispensing process for sampling of compounds with multiple nanoliters dispensing at the assay level, indicating in detail how we implemented the use of recycled PocketTips in our PerkinElmer EP3 workstations.