Establishment and optimization of NMR-based cell metabonomics study protocols for neonatal Sprague-Dawley rat cardiomyocytes

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• 作者：Ming Zhang^a ; ^b ; Bo Sun^b ; Qi Zhang^b ; Rong Gao^b ; Qiao Liu^b ; Fangting Dong^b ; Haiqin Fang^c ; Shuangqing Peng^c ; Famei Li^a ; ; Xianzhong Yan^b ; ^{yanxzhon@163.com}
• 关键词：Cell metabonomics ; Cellular metabolites extract ; NMR ; Cardiomyocytes
• 刊名：Analytical Biochemistry
• 出版年：2017
• 出版时间：15 January 2017
• 年：2017
• 卷：517
• 期：Complete
• 页码：50-52
• 全文大小：587 K
• 卷排序：517

文摘

A quenching, harvesting, and extraction protocol was optimized for cardiomyocytes NMR metabonomics analysis in this study. Trypsin treatment and direct scraping cells in acetonitrile were compared for sample harvesting. The results showed trypsin treatment cause normalized concentration increasing of phosphocholine and metabolites leakage, since the trypsin-induced membrane broken and long term harvesting procedures. Then the intracellular metabolite extraction efficiency of methanol and acetonitrile were compared. As a result, washing twice with phosphate buffer, direct scraping cells and extracting with acetonitrile were chosen to prepare cardiomyocytes extracts samples for metabonomics studies. This optimized protocol is rapid, effective, and exhibits greater metabolite retention.