Lysosomes are membrane-bound cell organelles that can be found in all mammalian cells. They contain digestive enzymes and are involved in multiple processes, including degradation of worn-out cell parts, antigen presentation and induction of programmed cell death. Mass spectrometry studies of lysosomes have been hampered by low sensitivity, since analysis of whole cell proteomes significantly reduce the detection of their relatively low abundant proteins. To overcome this issue, we applied 4 different commercial methods for the enrichment of lysosomes: gradient-based, filter-based, magnetic-bead based and subcellular fractionation, and performed quantitative proteomics analysis on these samples.