Macrophages are important immune cells operating at the forefront of innate immunity by taking up foreign particles and microbes through phagocytosis. The RAW 264.7 cell line is commonly used for experiments in the macrophage and phagocytosis field. However, little is known how its functions compare to primary macrophages. Here, we have performed an in-depth proteomics characterisation of phagosomes from RAW 264.7 and bone marrow-derived macrophages by quantifying more than 2500 phagosomal proteins. Our data indicates that there are significant differences for a large number of proteins including important receptors such as mannose receptor 1 and Siglec-1. Moreover, BMDM phagosomes mature considerably faster by fusion with endosomes and the lysosome when validated using fluorogenic phagocytic assays. We provide a valuable resource for researcher in the field and recommend careful use of the RAW 264.7 cell line when studying phagosome functions. This article is protected by copyright. All rights reserved.