Ishihama Yasushi, Sato Toshitaka, Tabata Tsuyoshi, Miyamoto Norimasa, Sagane Koji, Nagasu Takeshi, Oda Yoshiya
Nat Biotechnol. 2005 May;23(5):617-21. doi: 10.1038/nbt1086. Epub 2005 Apr 17.
An important challenge for proteomics is to be able to compare absolute protein levels across biological samples. Here we introduce an approach based on the use of culture-derived isotope tags (CDITs) for quantitative tissue proteome analysis. We cultured Neuro2A cells in a stable isotope-enriched medium and mixed them with mouse brain samples to serve as internal standards. Using CDITs, we identified and quantified a total of 1,000 proteins, 97-98% of which were expressed in both mouse whole brain and Neuro2A cells. CDITs also allow comprehensive and absolute protein quantification. Synthetic unlabeled peptides were used to quantify the corresponding proteins labeled with stable isotopes in Neuro2A cells, and the results were used to obtain the absolute amounts of 103 proteins in mouse whole brain. The expression levels correlated well with those in Neuro2A cells. Thus, the use of CDITs allows both relative and absolute quantitative proteome studies.
蛋白质组学面临的一项重要挑战是能够比较生物样品间的绝对蛋白质水平。在此，我们介绍一种基于使用培养衍生同位素标签（CDITs）进行定量组织蛋白质组分析的方法。我们在富含稳定同位素的培养基中培养Neuro2A细胞，并将其与小鼠脑样本混合作为内标。使用CDITs，我们共鉴定并定量了1000种蛋白质，其中97 - 98%在小鼠全脑和Neuro2A细胞中均有表达。CDITs还能实现全面且绝对的蛋白质定量。利用合成的未标记肽对Neuro2A细胞中用稳定同位素标记的相应蛋白质进行定量，并将结果用于获取小鼠全脑中103种蛋白质的绝对含量。其表达水平与Neuro2A细胞中的表达水平高度相关。因此，使用CDITs可进行相对和绝对定量蛋白质组研究。
