Individuality Normalization when Labeling with Isotopic Glycan Hydrazide Tags (INLIGHT): a novel glycan-relative quantification strategy.
Individuality Normalization when Labeling with Isotopic Glycan Hydrazide Tags (INLIGHT): a novel glycan-relative quantification strategy.
复制标题
DOI：
10.1007/s13361-013-0681-2
复制
发表时间：
2013-09
影响因子：
3.2
通讯作者：
Muddiman, David C.
中科院分区：
文献类型：
--
作者：
Walker, S. Hunter;Taylor, Amber D.;Muddiman, David C.
关键词：
提出了用于样品制备、数据分析和n -链聚糖相对定量的INLIGHT策略。甘聚糖用天然(L)或稳定同位素标记(H)肼试剂衍生化，并使用在线耦合Q萃取质谱仪的反相液相色谱法进行分析。一个简单的聚糖阶梯，麦芽糖糊精，首次用于演示相对定量策略的样品与可忽略不计的分析和生物变异性。结果表明，由于同位素重叠而进行了分子量校正，并对数据进行了采集后的归一化，以考虑系统变异性，实验H:L比与计算H:L比的图显示了以不同比例混合的麦芽糊精样品的一致性相关。我们还证明，INLIGHT方法可以量化离子丰度超过四个数量级的物种。INLIGHT策略在汇集的人体血浆中得到进一步验证，结果表明，采集后归一化比使用单个尖峰内标更有效。最后，当加入糖蛋白时，可以在复杂的生物基质中检测到糖基化的变化。糖蛋白峰值和检测聚糖水平变化的能力验证了样品制备和数据分析策略，使INLIGHT成为糖组学领域宝贵的相对量化策略。
The INLIGHT strategy for the sample preparation, data analysis, and relative quantification of N-linked glycans is presented. Glycans are derivatized with either natural (L) or stable-isotope labeled (H) hydrazide reagents and analyzed using reversed phase liquid chromatography coupled online to a Q Exactive mass spectrometer. A simple glycan ladder, maltodextrin, is first used to demonstrate the relative quantification strategy in samples with negligible analytical and biological variability. It is shown that after a molecular weight correction due to isotopic overlap and a post-acquisition normalization of the data to account for both the systematic variability, a plot of the experimental H:L ratio vs. the calculated H:L ratio exhibits a correlation of unity for maltodextrin samples mixed in different ratios. We also demonstrate that the INLIGHT approach can quantify species over four orders of magnitude in ion abundance. The INLIGHT strategy is further demonstrated in pooled human plasma, where it is shown that the post-acquisition normalization is more effective than using a single spiked-in internal standard. Finally, changes in glycosylation are able to be detected in complex biological matrices, when spiked with a glycoprotein. The ability to spike in a glycoprotein and detect change at the glycan level validates both the sample preparation and data analysis strategy, making INLIGHT an invaluable relative quantification strategy for the field of glycomics.
登录
查看更多内容
DOI：
10.1074/mcp.m111.011015
发表时间：
2011-09
期刊：
Molecular & cellular proteomics : MCP
影响因子：
--
作者：
Michalski A;Damoc E;Hauschild JP;Lange O;Wieghaus A;Makarov A;Nagaraj N;Cox J;Mann M;Horning S
通讯作者：
Horning S
DOI：
10.1016/j.jchromb.2009.02.040
发表时间：
2009-04-01
影响因子：
3
作者：
Andrews, Genna L.;Shuford, Christopher M.;Burnett, John C., Jr.;Hawkridge, Adam M.;Muddiman, David C.
通讯作者：
Muddiman, David C.
影响因子：
4
作者：
Kobata, A;Amano, J
通讯作者：
Amano, J
影响因子：
4.3
作者：
Alvarez-Manilla, Gerardo;Warren, Nicole Lynn;Orlando, Ron
通讯作者：
Orlando, Ron
DOI：
10.1007/s13361-011-0237-2
发表时间：
2011-12-01
影响因子：
3.2
作者：
Gokce, Emine;Shuford, Christopher M.;Muddiman, David C.
通讯作者：
Muddiman, David C.