基于准等重二甲基化标记的高通量蛋白质组选择性交叉标记装置

A cross-sequence labeling equipment for high-throughput proteome based on pseudo-isobaric dimethyl labeling

面对生物学及精准医学等领域多变量、大样本量的蛋白质组定量分析的需求,高通量的定量标记及分析已经成为近年来蛋白质组学方法发展的趋势。发展了一种基于准等重二甲基化标记策略的高通量肽段末端选择性交叉标记装置(pIDL-StageTip),借助简单的装置及离心力,有效地增加了定量标记的通量,并保证了肽段末端两步标记反应时间的可控性及操作的简便性。通过优化酸性条件下NaBD_3CN与NaBH_3CN体系的标记条件,得到了标准蛋白质酶解产物100%的标记效率、95%以上的标记选择性;在人源蛋白质组复杂体系下,标记效率大于99%,标记选择性为100%。基于该装置的定量方法具有很高的定量准确度及精密度。该装置为实现高可操作性、高准确度、高通量的蛋白质组定量标记提供了一个可靠的解决方案。

Facing the needs of multidimensional and multi-sample proteome quantification in the field of biology and precision medicine,high-throughput quantitative labeling and analysis have become the trend in recent years.A novel cross-sequence labeling equipment for high-throughput proteome based on pseudo-isobaric dimethyl labeling(pIDL-StageTip)was established.The equipment needs only simple devices and centrifugal force.The throughput of quantitative labeling is effectively increased,and the controllability of the two-step labeling reaction time at the terminals of the peptides and the ease of operation are ensured.By optimizing the labeling conditions of NaBDgCN and NaBHgCN under acidic conditions respectively,labeling efficiency of the standard proteolysis achieved 100%and labeling selectivity was more than 95%.Even in the complex human proteome,the labeling efficiency was greater than 99%and the labeling selectivity was 100%.The quantitative method based on the equipment showed high quantitative accuracy and precision.The equipment provides a reliable solution for quantitative proteome labeling with high operability,accuracy,and throughput.