FAQ

Last-modified: 2009-11-27 (¶â) 03:35:03 (3984d)

By testing the software, the selection of ¡Èset RT range (observable peptides)¡É seems to play a critical role in determining emPAI Calc output results.For example, using the same sample file provided for testing, the selection of ¡ÈShinoda et al (2006)¡É and ¡ÈNo retention time filtering¡É for emPAI calculation show quite different results. More elaborations are needed to explain how this factor affects the emPAI calculation.

  • The retention time range plays a critical role in determining the emPAI value because the RT filter determines the number of "observable" peptides. Briefly, we calculated the predicted RT of in silico peptides based on pre-defined equations; peptides out of the user-defined RT range are omitted from the calculation of "observable" peptides.

If well characterized RT (retention time) data or parameters are needed to calculate the emPAI values of a proteomics data set, the use of this program may be limited when such LC data or parameters are not available.

  • Basically, in LC-MS-based proteomic experiments, the RT is very robust parameter. Furthermore, our 4 RT filters (Shinoda et al., 2006; Ishihama, 2005; Krokhin et al., 2004; Petritis et al., 2003) covers a wide range of typical proteomics experiments. Thus, the use of this program is not highly limited. If emPAI Calc users adopt special LC conditions or LC parameters that are not available, the "No Retention Time filtering" option can be applied and the results are still a good indicator of protein abundance.

What is recommended MS instrument ?

QSTAR, LCQ, and LTQ.

The results are confused because for some proteins the number of the peptides that were actually observed for a protein is more than the number of peptides that were observable for the protein.

  • Selection of observable peptides is based on in silico tryptic digestion of a protein; it does not consider potential PTMs of a peptide. As for observed peptides, we count unique parent ions including different charge states from the same peptide sequence. Therefore, for some proteins, the number of observed- is greater than the number of observable peptides.