We are interested in the physical characterization of HIV-1 Virus-Like Particles (VLP) and purified capsid cores towards a better understanding of HIV-1 assembly. Here we focus on the modulation of their pleomorphic properties, i.e. the various global shapes of VLP or cores observed within common environmental and biological conditions. Using an original combination of biochemical approach, Atomic Force Microscopy (AFM) imaging and sophisticated image analysis, we quantitatively characterized the pleomorphism observed for HIV-1 VLP, as well as for purified viral cores. Analyzing three-dimensional high resolution AFM images of un-fixed VLP and viral cores, we conducted a complete morphological characterization of mature and immature particles at statistic levels. In addition, we revealed that the presence of the HIV-1 Psi-RNA inside viral cores is decisive for HIV-1 morphology. These findings shed light into the self-assembly and maturation of HIV-1 particle.