peer reviewed ; In Sub‐Saharan Africa, Anopheles gambiae Giles (Diptera: Culicidae) largely contributes to malaria transmission, in direct relation to environmental conditions influencing the vector ecology. Therefore, we carried out a proteomic analysis on An. gambiae sensu lato (s.l.) mosquitoes to compare their metabolic state, depending on different pesticide pressures by selecting areas with or without cotton crops, in two climatic regions. Adult mosquitoes were collected, and the proteomes were analysed by liquid chromatography with tandem mass spectrometry (LC‐MS/MS). The data are available via ProteomeXchange with identifier PXD016300. From a total of 1,182 identified proteins, 648 were retained for further statistical analysis and were attributed to biological functions, the most important of which is energy metabolism (120 proteins) followed by translation‐biogenesis (74), cytoskeleton (71), stress response (62), biosynthetic process (60), signalling (44), cellular respiration (38), cell redox homeostasis (25), DNA processing (17), pheromone binding (10), protein folding (9), RNA processing (9), other proteins (26) and unknown functions (83). The distribution of biological functions of all conditions was similar between climate zones or agricultural practices associated with different pesticide pressures. In the Sudano‐Sahelian region, 421 (91.3%) proteins were found in samples from areas both with and without cotton crops. By contrast, in the Sahelian region, only 271 (55.0%) were common to both crop areas, and 233 proteins were up‐regulated in samples from the cotton area. The focus was placed on differentially expressed proteins with putative roles in insecticide resistance, according to literature. This study provides the first whole‐body proteomic characterisation of An. gambiae s.l. in Burkina Faso, as a framework to strengthen vector control strategies and understand the environment‐vector interactions in different ecological sites.