ANALYTICAL METHOD FOR TEST SYSTEM FOR MULTIPLEX identification and quantification Measurement of the protein of interest in a biological sample CONTENT IN corresponding PROTEOTIPICHESKIH marker peptides

摘要

1. A process for preparing analytical test systems (MRM-test) for the multiplex identification and quantitative measurements of proteins of interest in a biological sample content corresponding proteotipicheskih marker peptides, comprising the following steps: 1) bioinformation unique identification marker for protein peptide sequences proteotipicheskih 2 ) selecting at least two marker peptide proteotipicheskih protein sequences, the most suitable method for the study monitor Yang multiple reactions, and 3) prediction of fragments of peptides, 4) prediction MRM-test in a list of marker peptides, their fragments, and the best parameters of detection by monitoring of multiple reactions, and 5) synthesis of one or more marker peptides; 6) determining the transition profile of single and multiple synthetic marker peptides by monitoring of multiple reactions under accelerated chromatographic gradient; 7) optimization MRM-test in accordance with the obtained profiles from a set of deletion peptide fragment cops, characterized by the lowest values ​​of the intensity in the mass spectrum; 8) purification of the peptides synthesized in the step 5), 9) the preparation of a biological sample, comprising removing major proteins; 10) protein identification in a biological sample with stab synthetic peptides by monitoring of multiple reactions under conditions normal chromatographic profiles on the basis of the gradient obtained in step 6); 11) determining the values ​​of retention time of marker peptides with entering set values ​​in MRM-testy.2.