Mass Spectrometry AMT approach
The AMT tag approach is based on advanced separations combined with very accurate MS measurements; in particular,
- ultra-high pressure capillary liquid chromatography (LC) combined with
- Fourier transform ion cyclotron resonance (FTICR) MS, and
- a supporting data analysis and management infrastructure.
Advantages include greater sensitivity and increased throughput. The greater sensitivity provides improved identification of lower abundance proteins, and thus better proteome coverage.
Basic schematic of the accurate mass and time (AMT) tag approach. The AMT approach consists of two main components:
- Creation of a mass and time (MT) tag database using a LC–MS/MS peptide identification strategy and
- use of LC-FTICR–MS for high-throughput accurate mass measurements which will be compared against the MT tag database for the identification of peptides and creation of an accurate mass and time (AMT) tag.
A peptide is validated as an AMT if:
- its observed mass, as measured by FTICR, agrees with the theoretical calculated mass of the PMT within 1 ppm MMA (Mass Measurement Accuracy).
- the LC elution times in the experiment wherein the PMT (Potential Mass Tag) was first identified has to agree with that observed in the FTICR experiment.
- the peptide must be unique within the annoteated genomic database (although distinctive elution time data should allow the disignation of AMTs having otherwise indistinguisable mases).
Only when these three cirteria are met, a peptide is designated as an AMT, and subsequently used to confidently identify a specific protein in subsequent proteome studies from the same species.
Without the need to reestablish the identiy of a peptide using time-consuming MS/MS analyses, multiple high-throughput studies to measure changes in relative protein abundances betweeen two or more different proteomes can be completed in rapid fashon based solely on the highly accurate maass measurements provided by FTICR.
References
- Jacobs JM, ME Monroe, W-J Qian, Y Shen, GA Anderson, and RD Smith. 2005. "Ultra-sensitive, high throughput and quantitative proteomics measurements." Int. J. Mass Spectrom. 240(3):195-212.
- Smith et al. 2002.