Rapid Proteomic Amelogenin Sex Estimation of Human and Cattle Remains Using Untargeted Evosep-timsTOF Mass Spectrometry

IF 1.7 3区化学 Q4 BIOCHEMICAL RESEARCH METHODS Rapid Communications in Mass Spectrometry Pub Date : 2025-03-11 DOI:10.1002/rcm.10022

Charllotte Blacka, Adam Dowle, Mik Lisowski, Michelle Alexander, Jessica Hendy, Kirsty Penkman, Jackie Mosely

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Abstract

Rationale

Sex estimation by analysis of amelogenin peptides in archaeological and fossil material has recently been gaining great traction within the fields of archaeology and palaeontology. Current widely used proteomic amelogenin sex estimation methods are hindered by relatively long mass spectrometric run times, or targeting peptides specific to human amelogenin proteins. Untargeted, high-throughput amelogenin sexing would be invaluable for a range of applications, from sex estimation of remains at mass grave sites to broadening the application of rapid amelogenin sexing to non-hominin species for husbandry and evolutionary studies.

Methods

A new acid etch protocol followed by Evosep-LC-TIMS-TOF mass spectrometry is presented for amelogenin analysis, providing global peptide data through rapid mass spectrometric methods in under 20 min per sample (including sample preparation, mass spectrometric acquisition and data processing). This sampling protocol was developed on modern cattle (Bos taurus) teeth, before Evosep-timsTOF partial validation with archaeological cattle and human (Homo sapiens) teeth, demonstrating the potential of straightforward application of this rapid amelogenin sexing method to a range of taxa.

Results

The rapid Evosep-LC-TIMS-TOF mass spectrometry methods gave comparable peptide counts to conventional long untargeted methods, while maintaining similar (or faster) acquisition times to those reported in methods targeting specific human amelogenin peptides. Implementation of the novel acid etch sampling approach also streamlined sample preparation without compromising peptide counts.

Conclusions

Rapid, untargeted Evosep-LC-TIMS-TOF mass spectrometry was successfully implemented in sex estimation of modern and archaeological material from Bos taurus and Homo sapiens teeth. This demonstrates an advancement in low-cost, high-throughput amelogenin sex estimation, for both human and zooarchaeological applications.

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非靶向Evosep-timsTOF质谱法快速测定人类和牛遗骸中的淀粉原蛋白性别

根据考古和化石材料中淀粉原蛋白肽的分析来估计性别最近在考古学和古生物学领域获得了很大的关注。目前广泛使用的蛋白质组学淀粉原蛋白性别估计方法受到相对较长的质谱运行时间或针对人类淀粉原蛋白特异性肽的阻碍。无目标、高通量的淀粉原蛋白性别鉴定在一系列应用中是无价的，从大规模墓地遗骸的性别估计到扩大淀粉原蛋白快速性别鉴定在畜牧业和进化研究中对非人类物种的应用。方法提出了一种新的酸蚀方法，采用Evosep-LC-TIMS-TOF质谱法分析淀粉原蛋白，通过快速质谱方法在20分钟内提供每个样品（包括样品制备，质谱采集和数据处理）的全局肽数据。在Evosep-timsTOF与考古牛和人类（智人）牙齿进行部分验证之前，该采样方案是在现代牛（Bos taurus）牙齿上开发的，证明了将这种快速变原蛋白性别测定方法直接应用于一系列分类群的潜力。结果快速的Evosep-LC-TIMS-TOF质谱方法提供的肽计数与传统的长非靶向方法相当，同时与针对特定人类淀粉原蛋白肽的方法保持相似（或更快）的获取时间。新型酸蚀取样方法的实施也简化了样品制备而不影响肽计数。结论快速、无靶向的Evosep-LC-TIMS-TOF质谱法可以成功地对现代和考古出土的金牛和智人牙齿材料进行性别鉴定。这表明在低成本、高通量的淀粉原性估计方面取得了进步，适用于人类和动物考古应用。

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来源期刊

Rapid Communications in Mass Spectrometry 化学-分析化学

CiteScore

4.10

自引率

5.00%

发文量

219

审稿时长

2.6 months

期刊介绍： Rapid Communications in Mass Spectrometry is a journal whose aim is the rapid publication of original research results and ideas on all aspects of the science of gas-phase ions; it covers all the associated scientific disciplines. There is no formal limit on paper length ("rapid" is not synonymous with "brief"), but papers should be of a length that is commensurate with the importance and complexity of the results being reported. Contributions may be theoretical or practical in nature; they may deal with methods, techniques and applications, or with the interpretation of results; they may cover any area in science that depends directly on measurements made upon gaseous ions or that is associated with such measurements.