Identification and characterization of the comprehensive bovine milk proteome has historically been limited due to the dichotomy of protein abundances within milk. The high abundance of a select few proteins, including caseins, α-lactalbumin, β-lactoglobulin, and serum albumin, has hindered intensive identification and characterization of the vast array of low abundance proteins within milk due to limitations in separation techniques and protein labeling capacity. In more recent years, the development and advancement of proteomics techniques has yielded valuable tools for characterization of the protein profile in bovine milk. More extensive fractionation and enrichment techniques, including the use of combinations of precipitation techniques, immunosorption, gel electrophoresis, chromatography, ultracentrifugation, and hexapeptide-based binding enrichment, have allowed for better isolation of lower abundance proteins for further downstream LC-MS/MS approaches. The different milk subfractions isolated during these processes can also be analyzed as individual entities to assess the protein profile unique to the different fractions, for instance, investigation of the skim milk-associated proteome versus the milk fat globule membrane-associated proteome. Updates to high-throughput methods, equipment, and software have also allowed for greater interpretation and visualization of the data. For instance, labeling techniques have enabled analysis of multiplexed samples and more accurate comparison of specific protein abundances and quantities across samples, while integration of gene ontology analysis has allowed for a more in-depth and visual representation of potential relationships between identified proteins. Inclusively, these developments in proteomic techniques have allowed for a rapid increase in the number of milk-associated proteins identified and a better grasp of the relationships and potential functionality of the proteins within the milk proteome.