In order to overcome the limitations of classic imaging in Histology during the actually era of multiomics, the multi-color "molecular microscope" by its emerging "molecular pictures" offers quantitative and spatial information about thousands of molecular profiles without labeling of potential targets. Healthy and diseased human tissues, as well as those of diverse invertebrate and vertebrate animal models, including genetically engineered species and cultured cells, can be easily analyzed by histology-directed MALDI imaging mass spectrometry. The aims of this review are to discuss a range of proteomic information emerging from MALDI mass spectrometry imaging comparative to classic histology, histochemistry and immunohistochemistry, with applications in biology and medicine, concerning the detection and distribution of structural proteins and biological active molecules, such as antimicrobial peptides and proteins, allergens, neurotransmitters and hormones, enzymes, growth factors, toxins and others. The molecular imaging is very well suited for discovery and validation of candidate protein biomarkers in neuroproteomics, oncoproteomics, aging and age-related diseases, parasitoproteomics, forensic, and ecotoxicology. Additionally, in situ proteome imaging may help to elucidate the physiological and pathological mechanisms involved in developmental biology, reproductive research, amyloidogenesis, tumorigenesis, wound healing, neural network regeneration, matrix mineralization, apoptosis and oxidative stress, pain tolerance, cell cycle and transformation under oncogenic stress, tumor heterogeneity, behavior and aggressiveness, drugs bioaccumulation and biotransformation, organism's reaction against environmental penetrating xenobiotics, immune signaling, assessment of integrity and functionality of tissue barriers, behavioral biology, and molecular origins of diseases. MALDI MSI is certainly a valuable tool for personalized medicine and "Eco-Evo-Devo" integrative biology in the current context of global environmental challenges.