The primary aim of the proposed research is to develop a fundamentally new proteomic tool, two-dimensional femtosecond laser-induced ionisation/dissociation tandem mass spectrometry (2D fs-LID MS/MS) and to use the power of this new proteomic approach to unveil the mechanisms of breast cancer endocrine therapy resistance and the role of ERalpha PTMs. The new MS platform will have similar advantages to other 2D spectroscopies (eg 2D NMR), namely peak correlations and spectral decongestion to grea tly aid molecular structural analysis and identification of protein PTMs. The applications of this new approach will benefit from exceptionally high level of protein sequence coverage and reliability of protein and PTM identifications. The 2D MS technology will be applied to in-depth characterisation of the role of PTMs in breast cancer endocrine resistance. The specific goals are: 1. Development of 2D tandem MS by combining linear ion trap femtosecond laser-induced ionisation/dissociation with covariance mapping to derive peptide fragment ion relationships of 1D tandem MS spectra and thus immensely facilitate interpretation of complex proteomic MS/MS data. 2. Application of the new technology to analysis of protein PTMs and their interplay that is key to understanding breast cancer endocrine therapy resistance and to development of efficient breast cancer therapies.