Wu Zhen, Xiang Weirong, Huang Lin, Li Shuwei, Zhang Xumin
State Key Laboratory of Genetic Engineering, Department of Biochemistry and Biophysics, School of Life Sciences, Fudan University, Shanghai 200438, China.
Nanjing Apollomics Biotech Inc., Nanjing, Jiangsu 210033, China.
Anal Chem. 2023 Mar 28;95(12):5169-5175. doi: 10.1021/acs.analchem.3c00237. Epub 2023 Mar 14.
Isobaric labeling has emerged as an indispensable quantitative proteomic approach for its unprecedented multiplexing capacity in a single analysis. Currently, different hyperplexing approaches have been developed to meet the demand for the increasing sample size in large-scale cohort analysis. In this report, we present a tribrid hyperplexing approach by the combinatorial use of three types of isobaric reagents, a novel isobaric tag 16-plex (IBT16) reagent and the widely used tandem mass tag (TMT; TMT11) and TMTpro (TMT18) reagents. After the determination of labeling efficiency and the optimization of testing conditions, we systematically evaluated the identification and quantification performance of the three labeling reagents in both independent and combinatorial manners using the mixtures of and HeLa peptides with different ratios. Our results reveal that the three reagents are quite similar in all testing aspects despite some differences, and the combination use of the three reagents could expand the multiplexing capacity to up to 45-plex. Furthermore, we conclude the advantages of IBT16 in the combination use and the preferred combinations for different practical applications. Data are available via ProteomeXchange with identifier PXD037498.
等压标记法因其在单次分析中具有前所未有的多重分析能力，已成为一种不可或缺的定量蛋白质组学方法。目前，已经开发出不同的超多重分析方法，以满足大规模队列分析中对不断增加的样本量的需求。在本报告中，我们通过组合使用三种类型的等压试剂、一种新型的16重等压标签（IBT16）试剂以及广泛使用的串联质量标签（TMT；TMT11）和TMTpro（TMT18）试剂，提出了一种三杂交超多重分析方法。在确定标记效率并优化测试条件后，我们使用不同比例的合成肽混合物和HeLa肽，以独立和组合的方式系统地评估了这三种标记试剂的鉴定和定量性能。我们的结果表明，尽管存在一些差异，但这三种试剂在所有测试方面都非常相似，并且三种试剂的组合使用可以将多重分析能力扩展到高达45重。此外，我们总结了IBT16在组合使用中的优势以及不同实际应用的优选组合。数据可通过ProteomeXchange获得，标识符为PXD037498。
