Methods and systems for experimental set-up and data analysis in targeted proteomics applications

Inventors

Escher, Claudia • Ossola, Reto • Rinner, Oliver • Reiter, Lukas

Assignees

Biognosys AG

Abstract

A method of the analysis of compounds with mass spectrometry and to instruments, substances, and methods for polypeptide analysis, in particular in targeted proteomics applications and based on indexed retention times as peptide specific property. The method of chemical analysis comprises the steps of: a) providing a first sample comprising peptides; b) performing LC-MS and determining the empirical retention time values of the peptides; c) translating the empirical retention time values into the indexed retention time scale and associating a reference indexed retention time value; d) providing a second complex sample comprising peptides; e) performing LC-MS and determining the empirical retention time values; f) translating the empirical retention time values of the peptides into the indexed retention time scale by numerically adapting the transformation function; and g) determining the predicted empirical retention time value of the peptides by using the numerically adapted transformation function determined in step f).

Core Innovation

The invention relates to a method of chemical analysis that uses LC-MS retention times and an indexed retention time scale to improve how peptides are identified, selected, and validated. A first complex sample contains a set of at least two reference peptides associated to an indexed retention time scale and at least one further peptide, and LC-MS is performed on the first complex sample to determine empirical retention time values (RTe) for the reference peptides and the further peptide.

The empirical retention time values (RTe) of the reference peptides are translated into the indexed retention time scale by associating each reference peptide with a reference indexed retention time value (iRTr) and associating the further peptide with an indexed retention time value (iRTa). A second complex sample is then provided comprising at least one polypeptide together with the set of at least two reference peptides, and LC-MS is performed on the second complex sample to determine empirical retention time values (RTe) of the reference peptides.

The empirical retention time values (RTe) of the reference peptides in the second complex sample are translated into the indexed retention time scale by numerically adapting a transformation function f so that the calculated indexed retention time values match the assigned reference indexed retention time values (iRTr). Using the numerically adapted transformation function f, a predicted empirical retention time value (RTp) is determined for the further peptide from its indexed retention time value (iRTa), and RTp is used for identification, determination of a retention time window, and definition of one or more regions where the further peptide cannot occur.

Claims Coverage

The claim coverage describes one independent chemical analysis method with three inventive features. It uses reference peptides to calibrate an indexed retention time scale, numerically adapts a transformation function from empirical retention times, and predicts RTp for further peptides for identification and retention-time constraints.

The inventive approach is based on calibrating an indexed retention time scale with reference peptides, adapting a transformation function from empirical retention times, and using RTp for peptide identification and retention-time window or exclusion-region definition.

Stated Advantages

Documented Applications