DNA甲基化是一种典型的表观遗传现象。已经开发了许多检测总体DNA甲基化水平的方法，其中从灵敏度，可再现性和成本的角度来看，LC-MS / MS已经成为一种极好的方法。然而，由于缺乏实验室测定所需的稳定性和标准化，LC-MS / MS方法具有局限性。本研究旨在建立一种可靠的测定方法，以确保对全球DNA甲基化水平进行高度准确的测量。所开发方法的至少三个方面。首先是发现使钠加合物减至最少的溶剂条件。第二个是使用以前的类似研究中未曾使用过的色谱柱，通过LC改进了核苷的分离。第三是成功减少了测量结果的不确定性，这是通过使用mdC的比例而不是在存在内标物的情况下使用绝对量进行校准而实现的。这些方面代表了优于先前报道的方法的优势。我们开发的方法能够在短时间内（8分钟）对DNA甲基化进行定量分析，并且以日间CV％小于5％表示的测量结果具有较高的可重复性。此外，通过测量培养的细胞系中总体DNA甲基化水平而获得的数据，无论是否进行药理学上的去甲基化处理，均支持将其用于生物医学研究。预期该测定法将使我们能够对各种细胞中的表观遗传学改变或畸变进行初步筛选。

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DNA methylation is a typical epigenetic phenomenon. Numerous methods for detecting global DNA methylation levels have been developed, among which LC-MS/MS has emerged as an excellent method from the viewpoint of sensitivity, reproducibility, and cost. However, LC-MS/MS methods have limitations due to a lack of the stability and the standardization required for a laboratory assay. The present study aimed to establish a robust assay that guarantees highly accurate measurements of global DNA methylation levels. There are at least three facets of the developed method. The first is discovery of the solvent conditions to minimize sodium adducts. The second is improvement of separation of nucleosides by LC using the columns that had not been used in previous similar studies. The third is success in reduction of the uncertainty of the measurement results, which was achieved by the calibration using the ratio of mdC but not the absolute amount in the presence of internal standards. These facets represent the advantage over methods reported previously. Our developed method enables quantification of DNA methylation with a short time length (8 min) for one analysis as well as with the high reproducibility of measurements that is represented by the inter-day CV% being less than 5%. In addition, data obtained from measuring global DNA methylation levels in cultured cell lines, with or without pharmacological demethylation, support its use for biomedical research. This assay is expected to allow us to conduct initial screening of epigenetic alterations or aberration in a variety of cells.