Key points are not available for this paper at this time.
LC-MS/MS has emerged as the method of choice for the identification and quantification of protein sample mixtures. For very complex samples such as complete proteomes, the most commonly used LC-MS/MS method, data-dependent acquisition (DDA) precursor selection, is of limited utility. The limited scan speed of current mass spectrometers along with the highly redundant selection of the most intense precursor ions generates a bias in the pool of identified proteins toward those of higher abundance. A directed LC-MS/MS approach that alleviates the limitations of DDA precursor ion selection by decoupling peak detection and sequencing of selected precursor ions is presented. In the first stage of the strategy, all detectable peptide ion signals are extracted from high resolution LC-MS feature maps or aligned sets of feature maps. The selected features or a subset thereof are subsequently sequenced in sequential, non-redundant directed LC-MS/MS experiments, and the MS/MS data are mapped back to the original LC-MS feature map in a fully automated manner. The strategy, implemented on an LTQ-FT MS platform, allowed the specific sequencing of 2,000 features per analysis and enabled the identification of more than 1,600 phosphorylation sites using a single reversed phase separation dimension without the need for time-consuming prefractionation steps. Compared with conventional DDA LC-MS/MS experiments, a substantially higher number of peptides could be identified from a sample, and this increase was more pronounced for low intensity precursor ions. LC-MS/MS has emerged as the method of choice for the identification and quantification of protein sample mixtures. For very complex samples such as complete proteomes, the most commonly used LC-MS/MS method, data-dependent acquisition (DDA) precursor selection, is of limited utility. The limited scan speed of current mass spectrometers along with the highly redundant selection of the most intense precursor ions generates a bias in the pool of identified proteins toward those of higher abundance. A directed LC-MS/MS approach that alleviates the limitations of DDA precursor ion selection by decoupling peak detection and sequencing of selected precursor ions is presented. In the first stage of the strategy, all detectable peptide ion signals are extracted from high resolution LC-MS feature maps or aligned sets of feature maps. The selected features or a subset thereof are subsequently sequenced in sequential, non-redundant directed LC-MS/MS experiments, and the MS/MS data are mapped back to the original LC-MS feature map in a fully automated manner. The strategy, implemented on an LTQ-FT MS platform, allowed the specific sequencing of 2,000 features per analysis and enabled the identification of more than 1,600 phosphorylation sites using a single reversed phase separation dimension without the need for time-consuming prefractionation steps. Compared with conventional DDA LC-MS/MS experiments, a substantially higher number of peptides could be identified from a sample, and this increase was more pronounced for low intensity precursor ions. Over the past decade, MS has emerged as the method of choice for the identification and quantification of proteins in very complex biological samples (1Aebersold R. Mann M. Mass spectrometry-based proteomics.Nature. 2003; 422: 198-207Crossref PubMed Scopus (5540) Google Scholar). In the most widely used implementation, referred to as shotgun proteomics, protein samples are first digested, and the resulting peptide mixtures are then chromatographically separated and finally sequenced by automated MS/MS. Because of its conceptual and experimental simplicity, the shotgun approach has become a very popular method for the identification of proteins in a wide range of biological samples and, in combination with stable isotope labeling, for quantitative proteomics studies (2Gygi S.P. Rist B. Gerber S.A. Turecek F. Gelb M.H. Aebersold R. Quantitative analysis of complex protein mixtures using isotope-coded affinity tags.Nat. Biotechnol. 1999; 17: 994-999Crossref PubMed Scopus (4321) Google Scholar, 3Ong S.E. Kratchmarova I. Mann M. Properties of 13C-substituted arginine in stable isotope labeling by amino acids in cell culture (SILAC).J. Proteome Res. 2003; 2: 173-181Crossref PubMed Scopus (370) Google Scholar, 4Schmidt A. Kellermann J. Lottspeich F. A novel strategy for quantitative proteomics using isotope-coded protein labels.Proteomics. 2005; 5: 4-15Crossref PubMed Scopus (432) Google Scholar). Recent technical improvements in MS instrumentation, database searching, and result validation as well as advances in database annotation now make it possible to routinely identify hundreds to a few thousands of proteins in complex biological samples (5Brunner E. Ahrens C.H. Mohanty S. Baetschmann H. Loevenich S. Potthast F. Deutsch E.W. Panse C. de Lichtenberg U. H. J. S. J. F. E. R. Aebersold R. A of the Biotechnol. PubMed Scopus Google Scholar, B. A. C. A. J. B. A. of and protein in and PubMed Scopus Google Scholar, J. C. Mann M. The more than a of PubMed Scopus Google Scholar, de Mann M. of complete analysis by mass as a PubMed Scopus Google this shotgun proteomics is of and the protein of low (5Brunner E. Ahrens C.H. Mohanty S. Baetschmann H. Loevenich S. Potthast F. Deutsch E.W. Panse C. de Lichtenberg U. H. J. S. J. F. E. R. Aebersold R. A of the Biotechnol. PubMed Scopus Google Scholar, de Mann M. of complete analysis by mass as a PubMed Scopus Google Scholar). is a of limited sequencing speed of current that are of precursor ion in complex samples with the redundant selection of a subset of precursor ions are in of the sample identification of the low intensity is B. M. Aebersold R. with peptide 2005; PubMed Scopus Google Scholar, J. Ahrens C.H. S. an for validation and of mass PubMed Scopus Google Scholar, H. A for and of protein in shotgun PubMed Scopus Google to on data-dependent acquisition (DDA) used data-dependent precursor reversed used data-dependent precursor reversed precursor ion selection, directed peptide sequencing the of the MS/MS analysis on non-redundant and precursor the analysis and the of analysis B. Aebersold R. Mass and protein PubMed Scopus Google Scholar, M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google Scholar). In this a strategy by all features that peptides are extracted from LC-MS maps and subsequently to in to the identification of all detectable B. S. of proteomics for and Proteome Res. PubMed Scopus Google Scholar). Because the acquisition of and is in this is well for directed sequencing and has to or of proteins B. A. B. C. A. Kellermann J. F. F. Lottspeich F. Quantitative of the in a 5: PubMed Scopus Google Scholar, A. I. H. Aebersold R. analysis by mass 2003; PubMed Scopus Google is to has the to higher sequencing speed in with Because the peptides are on the sample of the sample are the first to the features and for directed the decoupling of feature detection and sequencing highly and high mass A directed sequencing strategy has on high mass to peptides of single proteins in complex mixtures S. M. S.A. B. A. M. S.A. identification of PubMed Scopus Google and for the detection of low peptide Aebersold R. B. The of for shotgun PubMed Scopus Google Scholar). the number of peptide ion was limited to a few per a number of sequencing than are of in DDA The number of could be by sample and LC-MS/MS of the the a high mass that of mass by was used for directed the and of the and was of the number of possible to in a The strategy was by to peptide features from maps and to features LC-MS/MS M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google from the identified H. I. R. and in H. I. R. and in directed sequencing of the features on the by LC-MS/MS of the sample, and map the MS/MS data back to the feature The of the directed MS/MS approach was by of complex peptide and mixtures from The data the high and of the method to identify a higher number of peptides from the sample in a number of LC-MS/MS with DDA LC-MS/MS in the of low intensity precursor a and and mass method for directed sequencing of a high number of peptide ions complex mixtures. In to and data acquisition DDA the directed approach and In a first peptide ion signals are extracted from the in LC-MS/MS For this the and In a such features to directed sequencing in LC-MS/MS and the identified features mapped back to the of make the for this method and it data R. M. Deutsch E.W. B. B. E. R. H. S. E. R. R. Aebersold R. A of mass data and its to proteomics Biotechnol. PubMed Scopus Google Scholar, A. E. Aebersold R. A for proteins by mass 2003; PubMed Scopus Google Scholar, A. E. Aebersold R. to the of peptide by MS/MS and database PubMed Scopus Google for for MS are Proteome the approach is to high MS that sequencing by is to that high resolution are for data acquisition and for MS the LTQ-FT used in this are the of and data be in the LC-MS/MS with of the speed and for high maps. Compared with current DDA the directed strategy the feature the identification of peptides with low intensity precursor ion peak selection in DDA LC-MS/MS analysis is on single peptide ion detection all signals of an peptide the selection of peptide ions on the of as well as the of and precursor the of peptide ions to signals and and the of a feature with a high of peptide ions. the directed sequencing of features in a higher number of peptides identified with by is that the number of by the directed approach was than by DDA the for data in directed the be and to the feature For a higher number of very intense features could be identified in LC-MS/MS by ion and scan and scan used for low peptide ions. is to that the directed approach the of all features that could be to a peptide in the first analysis using MS by the in this more than phosphorylation sites by analysis of that a peak could be identified on the of or using the for MS/MS data acquisition J. A method for analysis by ion mass PubMed Scopus Google Scholar). The of for feature for the identification of more peptides be used to peptide ions with the directed approach the of for precursor ions with specific the of a peptide that is most to an ion stable isotope labeling M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google Scholar, B. A. B. C. A. Kellermann J. F. F. Lottspeich F. Quantitative of the in a 5: PubMed Scopus Google Scholar, A. I. H. Aebersold R. analysis by mass 2003; PubMed Scopus Google Scholar, A. of for proteins by shotgun 2005; PubMed Scopus Google isotope by selected with Rist B. Aebersold R. identification with a single mass of a peptide and database PubMed Scopus Google of peptides J. M. A. M. Aebersold R. of peptides from 5: PubMed Scopus Google and redundant identification of peptides of samples M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google complex the most of directed DDA LC-MS/MS from the that the directed approach with the of LC-MS/MS analysis that all precursor ions in the be sequenced using LC-MS/MS de Mann M. of complete analysis by mass as a PubMed Scopus Google Scholar, B. M. Aebersold R. with peptide 2005; PubMed Scopus Google Scholar, J. Ahrens C.H. S. an for validation and of mass PubMed Scopus Google Scholar, H. A for and of protein in shotgun PubMed Scopus Google Scholar). in the identification of peptides of high using the directed the sequencing speed of the MS used is a with to the detectable peptide ion in a sample to be a higher number of peptides identified by using the approach with most a precursor intensity than that of the of peptides by the DDA The identification of low intensity peptide ions by directed LC-MS/MS was pronounced for of peptide sequencing by DDA LC-MS/MS of the limited MS sequencing speed and a more sequencing of precursor ions by the directed the number of peptides identified by directed LC-MS/MS with be to the that very low intensity the detection in all higher mass and are to and to maps. A higher range in the be this is limited by the of the cell mass the combination of and in a ion mass PubMed Scopus Google Scholar, C. R. Mass by in mass Mass PubMed Scopus Google Scholar). the intensity range of peptide ions be by time-consuming sample (5Brunner E. Ahrens C.H. Mohanty S. Baetschmann H. Loevenich S. Potthast F. Deutsch E.W. Panse C. de Lichtenberg U. H. J. S. J. F. E. R. Aebersold R. A of the Biotechnol. PubMed Scopus Google Scholar, M. H. Aebersold R. complete by PubMed Scopus Google be used in combination with the directed feature to be on by the MS to the MS detection this for a of low features scan was improvements in peak detection of precursor ions could increase the number of for low peptide directed sequencing of high features the identification of a higher number of peptides with than current DDA For more than 1,600 phosphorylation sites could be identified using a single dimension reversed phase separation without the need for time-consuming sample prefractionation steps. The implemented used are and with data and to most high LC-MS the of high LC-MS/MS and its to identify features of and a wide intensity the directed approach is well for and high of complex protein samples and wide in Over the past decade, MS has emerged as the method of choice for the identification and quantification of proteins in very complex biological samples (1Aebersold R. Mann M. Mass spectrometry-based proteomics.Nature. 2003; 422: 198-207Crossref PubMed Scopus (5540) Google Scholar). In the most widely used implementation, referred to as shotgun proteomics, protein samples are first digested, and the resulting peptide mixtures are then chromatographically separated and finally sequenced by automated MS/MS. Because of its conceptual and experimental simplicity, the shotgun approach has become a very popular method for the identification of proteins in a wide range of biological samples and, in combination with stable isotope labeling, for quantitative proteomics studies (2Gygi S.P. Rist B. Gerber S.A. Turecek F. Gelb M.H. Aebersold R. Quantitative analysis of complex protein mixtures using isotope-coded affinity tags.Nat. Biotechnol. 1999; 17: 994-999Crossref PubMed Scopus (4321) Google Scholar, 3Ong S.E. Kratchmarova I. Mann M. Properties of 13C-substituted arginine in stable isotope labeling by amino acids in cell culture (SILAC).J. Proteome Res. 2003; 2: 173-181Crossref PubMed Scopus (370) Google Scholar, 4Schmidt A. Kellermann J. Lottspeich F. A novel strategy for quantitative proteomics using isotope-coded protein labels.Proteomics. 2005; 5: 4-15Crossref PubMed Scopus (432) Google Scholar). Recent technical improvements in MS instrumentation, database searching, and result validation as well as advances in database annotation now make it possible to routinely identify hundreds to a few thousands of proteins in complex biological samples (5Brunner E. Ahrens C.H. Mohanty S. Baetschmann H. Loevenich S. Potthast F. Deutsch E.W. Panse C. de Lichtenberg U. H. J. S. J. F. E. R. Aebersold R. A of the Biotechnol. PubMed Scopus Google Scholar, B. A. C. A. J. B. A. of and protein in and PubMed Scopus Google Scholar, J. C. Mann M. The more than a of PubMed Scopus Google Scholar, de Mann M. of complete analysis by mass as a PubMed Scopus Google Scholar). this shotgun proteomics is of and the protein of low (5Brunner E. Ahrens C.H. Mohanty S. Baetschmann H. Loevenich S. Potthast F. Deutsch E.W. Panse C. de Lichtenberg U. H. J. S. J. F. E. R. Aebersold R. A of the Biotechnol. PubMed Scopus Google Scholar, de Mann M. of complete analysis by mass as a PubMed Scopus Google Scholar). is a of limited sequencing speed of current that are of precursor ion in complex samples with the redundant selection of a subset of precursor ions are in of the sample identification of the low intensity is B. M. Aebersold R. with peptide 2005; PubMed Scopus Google Scholar, J. Ahrens C.H. S. an for validation and of mass PubMed Scopus Google Scholar, H. A for and of protein in shotgun PubMed Scopus Google Scholar). In to on data-dependent acquisition (DDA) used data-dependent precursor reversed used data-dependent precursor reversed precursor ion selection, directed peptide sequencing the of the MS/MS analysis on non-redundant and precursor the analysis and the of analysis B. Aebersold R. Mass and protein PubMed Scopus Google Scholar, M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google Scholar). In this a strategy by all features that peptides are extracted from LC-MS maps and subsequently to in to the identification of all detectable B. S. of proteomics for and Proteome Res. PubMed Scopus Google Scholar). Because the acquisition of and is in this is well for directed sequencing and has to or of proteins B. A. B. C. A. Kellermann J. F. F. Lottspeich F. Quantitative of the in a 5: PubMed Scopus Google Scholar, A. I. H. Aebersold R. analysis by mass 2003; PubMed Scopus Google Scholar). The is to has the to higher sequencing speed in with Because the peptides are on the sample of the sample are the first to the features and for directed the decoupling of feature detection and sequencing highly and high mass A directed sequencing strategy has on high mass to peptides of single proteins in complex mixtures S. M. S.A. B. A. M. S.A. identification of PubMed Scopus Google and for the detection of low peptide Aebersold R. B. The of for shotgun PubMed Scopus Google Scholar). the number of peptide ion was limited to a few per a number of sequencing than are of in DDA The number of could be by sample and LC-MS/MS of the In the a high mass that of mass by was used for directed the and of the and was of the number of possible to in a The strategy was by to peptide features from maps and to features LC-MS/MS M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google from the identified H. I. R. and in H. I. R. and in directed sequencing of the features on the by LC-MS/MS of the sample, and map the MS/MS data back to the feature The of the directed MS/MS approach was by of complex peptide and mixtures from The data the high and of the method to identify a higher number of peptides from the sample in a number of LC-MS/MS with DDA LC-MS/MS in the of low intensity precursor ions. a and and mass method for directed sequencing of a high number of peptide ions complex mixtures. In to and data acquisition DDA the directed approach and In a first peptide ion signals are extracted from the in LC-MS/MS For this the and In a such features to directed sequencing in LC-MS/MS and the identified features mapped back to the of make the for this method and it data R. M. Deutsch E.W. B. B. E. R. H. S. E. R. R. Aebersold R. A of mass data and its to proteomics Biotechnol. PubMed Scopus Google Scholar, A. E. Aebersold R. A for proteins by mass 2003; PubMed Scopus Google Scholar, A. E. Aebersold R. to the of peptide by MS/MS and database PubMed Scopus Google for for MS are Proteome the approach is to high MS that sequencing by is to that high resolution are for data acquisition and for MS the LTQ-FT used in this are the of and data be in the LC-MS/MS with of the speed and for high maps. Compared with current DDA the directed strategy the feature the identification of peptides with low intensity precursor ion peak selection in DDA LC-MS/MS analysis is on single peptide ion detection all signals of an peptide the selection of peptide ions on the of as well as the of and precursor the of peptide ions to signals and and the of a feature with a high of peptide ions. the directed sequencing of features in a higher number of peptides identified with by is that the number of by the directed approach was than by DDA the for data in directed the be and to the feature For a higher number of very intense features could be identified in LC-MS/MS by ion and scan and scan used for low peptide ions. is to that the directed approach the of all features that could be to a peptide in the first analysis using MS by the in this more than phosphorylation sites by analysis of that a peak could be identified on the of or using the for MS/MS data acquisition J. A method for analysis by ion mass PubMed Scopus Google Scholar). The of for feature for the identification of more peptides be used to peptide ions with the directed approach the of for precursor ions with specific the of a peptide that is most to an ion stable isotope labeling M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google Scholar, B. A. B. C. A. Kellermann J. F. F. Lottspeich F. Quantitative of the in a 5: PubMed Scopus Google Scholar, A. I. H. Aebersold R. analysis by mass 2003; PubMed Scopus Google Scholar, A. of for proteins by shotgun 2005; PubMed Scopus Google isotope by selected with Rist B. Aebersold R. identification with a single mass of a peptide and database PubMed Scopus Google of peptides J. M. A. M. Aebersold R. of peptides from 5: PubMed Scopus Google and redundant identification of peptides of samples M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google complex the most of directed DDA LC-MS/MS from the that the directed approach with the of LC-MS/MS analysis that all precursor ions in the be sequenced using LC-MS/MS de Mann M. of complete analysis by mass as a PubMed Scopus Google Scholar, B. M. Aebersold R. with peptide 2005; PubMed Scopus Google Scholar, J. Ahrens C.H. S. an for validation and of mass PubMed Scopus Google Scholar, H. A for and of protein in shotgun PubMed Scopus Google Scholar). in the identification of peptides of high using the directed the sequencing speed of the MS used is a with to the detectable peptide ion in a sample to be a higher number of peptides identified by using the approach with most a precursor intensity than that of the of peptides by the DDA The identification of low intensity peptide ions by directed LC-MS/MS was pronounced for of peptide sequencing by DDA LC-MS/MS of the limited MS sequencing speed and a more sequencing of precursor ions by the directed the number of peptides identified by directed LC-MS/MS with be to the that very low intensity the detection in all higher mass and are to and to maps. A higher range in the be this is limited by the of the cell mass the combination of and in a ion mass PubMed Scopus Google Scholar, C. R. Mass by in mass Mass PubMed Scopus Google Scholar). the intensity range of peptide ions be by time-consuming sample (5Brunner E. Ahrens C.H. Mohanty S. Baetschmann H. Loevenich S. Potthast F. Deutsch E.W. Panse C. de Lichtenberg U. H. J. S. J. F. E. R. Aebersold R. A of the Biotechnol. PubMed Scopus Google Scholar, M. H. Aebersold R. complete by PubMed Scopus Google be used in combination with the directed feature to be on by the MS to the MS detection this for a of low features scan was improvements in peak detection of precursor ions could increase the number of for low peptide directed sequencing of high features the identification of a higher number of peptides with than current DDA For more than 1,600 phosphorylation sites could be identified using a single dimension reversed phase separation without the need for time-consuming sample prefractionation steps. The implemented used are and with data and to most high LC-MS the of high LC-MS/MS and its to identify features of and a wide intensity the directed approach is well for and high of complex protein samples and wide in a and and mass method for directed sequencing of a high number of peptide ions complex mixtures. In to and data acquisition DDA the directed approach and In a first peptide ion signals are extracted from the in LC-MS/MS For this the and In a such features to directed sequencing in LC-MS/MS and the identified features mapped back to the of make the for this method and it data R. M. Deutsch E.W. B. B. E. R. H. S. E. R. R. Aebersold R. A of mass data and its to proteomics Biotechnol. PubMed Scopus Google Scholar, A. E. Aebersold R. A for proteins by mass 2003; PubMed Scopus Google Scholar, A. E. Aebersold R. to the of peptide by MS/MS and database PubMed Scopus Google for for MS are Proteome the approach is to high MS that sequencing by is to that high resolution are for data acquisition and for MS the LTQ-FT used in this are the of and data be in the LC-MS/MS with of the speed and for high maps. Compared with current DDA the directed strategy the feature the identification of peptides with low intensity precursor ion peak selection in DDA LC-MS/MS analysis is on single peptide ion detection all signals of an peptide the selection of peptide ions on the of as well as the of and precursor the of peptide ions to signals and and the of a feature with a high of peptide ions. the directed sequencing of features in a higher number of peptides identified with by is that the number of by the directed approach was than by DDA the for data in directed the be and to the feature For a higher number of very intense features could be identified in LC-MS/MS by ion and scan and scan used for low peptide ions. is to that the directed approach the of all features that could be to a peptide in the first analysis using MS by the in this more than phosphorylation sites by analysis of that a peak could be identified on the of or using the for MS/MS data acquisition J. A method for analysis by ion mass PubMed Scopus Google Scholar). The of for feature for the identification of more peptides be used to peptide ions with the directed approach the of for precursor ions with specific the of a peptide that is most to an ion stable isotope labeling M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google Scholar, B. A. B. C. A. Kellermann J. F. F. Lottspeich F. Quantitative of the in a 5: PubMed Scopus Google Scholar, A. I. H. Aebersold R. analysis by mass 2003; PubMed Scopus Google Scholar, A. of for proteins by shotgun 2005; PubMed Scopus Google isotope by selected with Rist B. Aebersold R. identification with a single mass of a peptide and database PubMed Scopus Google of peptides J. M. A. M. Aebersold R. of peptides from 5: PubMed Scopus Google and redundant identification of peptides of samples M. M. M. Aebersold R. mass and for the analysis of protein Biotechnol. PubMed Scopus Google Scholar). For complex the most of directed DDA LC-MS/MS from the that the directed approach with the of LC-MS/MS analysis that all precursor ions in the be sequenced using LC-MS/MS de Mann M. of complete analysis by mass as a PubMed Scopus Google Scholar, B. M. Aebersold R. with peptide 2005; PubMed Scopus Google Scholar, J. Ahrens C.H. S. an for validation and of mass PubMed Scopus Google Scholar, H. A for and of protein in shotgun PubMed Scopus Google Scholar). in the identification of peptides of high using the directed the sequencing speed of the MS used is a with to the detectable peptide ion in a sample to be a higher number of peptides identified by using the approach with most a precursor intensity than that of the of peptides by the DDA The identification of low intensity peptide ions by directed LC-MS/MS was pronounced for of peptide sequencing by DDA LC-MS/MS of the limited MS sequencing speed and a more sequencing of precursor ions by the directed the number of peptides identified by directed LC-MS/MS with be to the that very low intensity the detection in all higher mass and are to and to maps. A higher range in the be this is limited by the of the cell mass the combination of and in a ion mass PubMed Scopus Google Scholar, C. R. Mass by in mass Mass PubMed Scopus Google Scholar). the intensity range of peptide ions be by time-consuming sample (5Brunner E. Ahrens C.H. Mohanty S. Baetschmann H. Loevenich S. Potthast F. Deutsch E.W. Panse C. de Lichtenberg U. H. J. S. J. F. E. R. Aebersold R. A of the Biotechnol. PubMed Scopus Google Scholar, M. H. Aebersold R. complete by PubMed Scopus Google be used in combination with the directed feature to be on by the MS to the MS detection this for a of low features scan was improvements in peak detection of precursor ions could increase the number of for low peptide ions. In directed sequencing of high features the identification of a higher number of peptides with than current DDA For more than 1,600 phosphorylation sites could be identified using a single dimension reversed phase separation without the need for time-consuming sample prefractionation steps. The implemented used are and with data and to most high LC-MS the of high LC-MS/MS and its to identify features of and a wide intensity the directed approach is well for and high of complex protein samples and wide in and for with with
Schmidt et al. (Thu,) studied this question.