Mehl John T, Cummings John J, Rohde Ellen, Yates Nathan N
Molecular Profiling, Merck Research Laboratories, Rahway, NJ 07065, USA.
Rapid Commun Mass Spectrom. 2003;17(14):1600-10. doi: 10.1002/rcm.1092.
Atmospheric-pressure matrix-assisted laser desorption/ionization (AP-MALDI) ion trap mass spectrometry (ITMS) has been evaluated for automated protein identification. By using signal averaging and long ion-injection times, protein identification limits in the 50-fmol range are achieved for standard protein digests. Data acquisition requires 7.5 min or less per sample and the MS/MS spectra files are automatically processed using the SEQUEST database searching algorithm. AP-MALDI-ITMS was compared with the widely used methods of microLC/MS/MS (ion trap) and automated MALDI-TOF peptide mass mapping. Sample throughput is 10-fold greater using AP-MALDI compared with microcapillary liquid chromatography/tandem mass spectrometry (microLC/MS/MS). The protein sequence coverage obtained from AP-MALDI-MS/MS spectra matched by SEQUEST is lower compared with microLC/MS/MS and MALDI-TOF mass mapping. However, by using the AP-MALDI full-scan peptide mass fingerprint spectrum, sequence coverage is increased. AP-MALDI-ITMS was applied for the analysis of Coomassie blue stained gels and was found to be a useful platform for rapid protein identification.
常压基质辅助激光解吸/电离（AP-MALDI）离子阱质谱（ITMS）已用于蛋白质自动鉴定评估。通过信号平均和长离子注入时间，标准蛋白质消化物的蛋白质鉴定限可达50飞摩尔范围。每个样品的数据采集时间为7.5分钟或更短，并且使用SEQUEST数据库搜索算法自动处理MS/MS光谱文件。将AP-MALDI-ITMS与广泛使用的微液相色谱/串联质谱（离子阱）和自动MALDI-TOF肽质量图谱方法进行了比较。与微毛细管液相色谱/串联质谱（microLC/MS/MS）相比，使用AP-MALDI时样品通量提高了10倍。通过SEQUEST匹配的AP-MALDI-MS/MS光谱获得的蛋白质序列覆盖率低于microLC/MS/MS和MALDI-TOF质量图谱。然而，通过使用AP-MALDI全扫描肽质量指纹图谱，序列覆盖率得以提高。AP-MALDI-ITMS用于考马斯亮蓝染色凝胶的分析，被发现是快速蛋白质鉴定的有用平台。
