Mass spectrometry (MS) has played an increasingly prominent role in proteomics and structure biology because it shows superior capabilities in identification, quantification and structural characterization of proteins. To realize its full potential in protein analysis, significant progress has been made in developing innovative techniques and reagents that can couple to MS detection. This dissertation demonstrates the use of polymeric supramolecular assemblies for enhanced protein detection in complex biological mixtures by MS. An amphiphilic random co-polymer scaffold is developed to form functional supramolecular assemblies for protein/ peptide enrichment. The influences of charge density and functional group pKa on host-guest interactions within the assemblies are fundamentally investigated. In practice, these new materials enable specific isolation of target peptides from complex mixtures, as well as enhance MS detection/ quantification of protein biomarker in human breast milk. In parallel to protein detection, this dissertation also describes the development of a series of small-molecule covalent labeling (CL) reagents that are capable of studying protein higher order structure and protein-protein interactions when coupled with MS.