This review discusses selected reaction monitoring (SRM) as a method for quantitative proteomics. SRM is a technique that allows precise quantification of low-abundance proteins in complex mixtures, complementing shotgun proteomics approaches. In SRM, a predefined precursor ion and one of its fragment ions are selected by a triple quadrupole mass spectrometer and monitored over time. A series of transitions (precursor/fragment ion pairs) in combination with retention time can constitute a definitive assay. SRM offers high selectivity, sensitivity, and dynamic range, making it ideal for accurate quantification of predefined protein sets in systems biology. The review outlines the steps for establishing an SRM experiment, including the selection of a target protein set, peptide selection, and the optimization and validation of transitions. It discusses factors affecting SRM performance, such as ionization and fragmentation conditions, and the importance of selecting peptides that uniquely identify the target protein. Post-translational modifications and chemically induced modifications are also considered, as they can affect SRM results. The review also covers the validation of transitions, which is crucial to ensure that the quantified signals indeed derive from the targeted peptide. This can be done by acquiring MS/MS spectra and comparing them with predicted peptide fragments. Additionally, the use of heavy isotope-labelled peptides can help in validating transitions when not available. The review highlights the importance of normalization in SRM experiments to account for variations in sample processing and background. It discusses both label-free and isotope-based quantification methods, with isotope-based methods offering higher precision. The review also addresses the challenges of sample amount normalization and the importance of using invariant proteins for normalization. Finally, the review discusses absolute quantification, which requires the addition of isotopically labelled reference peptides or proteins to the sample. This allows for the precise determination of protein abundance, such as copy numbers per cell or protein concentration in blood. The review concludes that SRM is a powerful tool for quantitative proteomics, offering high sensitivity, selectivity, and accuracy.This review discusses selected reaction monitoring (SRM) as a method for quantitative proteomics. SRM is a technique that allows precise quantification of low-abundance proteins in complex mixtures, complementing shotgun proteomics approaches. In SRM, a predefined precursor ion and one of its fragment ions are selected by a triple quadrupole mass spectrometer and monitored over time. A series of transitions (precursor/fragment ion pairs) in combination with retention time can constitute a definitive assay. SRM offers high selectivity, sensitivity, and dynamic range, making it ideal for accurate quantification of predefined protein sets in systems biology. The review outlines the steps for establishing an SRM experiment, including the selection of a target protein set, peptide selection, and the optimization and validation of transitions. It discusses factors affecting SRM performance, such as ionization and fragmentation conditions, and the importance of selecting peptides that uniquely identify the target protein. Post-translational modifications and chemically induced modifications are also considered, as they can affect SRM results. The review also covers the validation of transitions, which is crucial to ensure that the quantified signals indeed derive from the targeted peptide. This can be done by acquiring MS/MS spectra and comparing them with predicted peptide fragments. Additionally, the use of heavy isotope-labelled peptides can help in validating transitions when not available. The review highlights the importance of normalization in SRM experiments to account for variations in sample processing and background. It discusses both label-free and isotope-based quantification methods, with isotope-based methods offering higher precision. The review also addresses the challenges of sample amount normalization and the importance of using invariant proteins for normalization. Finally, the review discusses absolute quantification, which requires the addition of isotopically labelled reference peptides or proteins to the sample. This allows for the precise determination of protein abundance, such as copy numbers per cell or protein concentration in blood. The review concludes that SRM is a powerful tool for quantitative proteomics, offering high sensitivity, selectivity, and accuracy.