Protein ubiquitination is essential for multiple physiological processes through regulating the stability or function of target proteins and has been found to play critical roles in human cancers. However, the protein ubiquitination profile of human metastatic colon adenocarcinoma tissue has not been elucidated yet. In this study, a proprietary ubiquitin branch (K-ε-GG) antibody-based label-free quantitative proteomics and bioinformatics were performed to identify the global protein ubiquitination profile between human primary (Colon) and metastatic colon adenocarcinoma (Meta) tissues.