Cell cultures suspended in bioreactors in a fluid environment are the basis for cell expansion and important medical products manufacturing. Assessing local cell distribution within bioreactors may provide information to increase cell production efficiency. Hydrodynamics characterizations of bioreactors are typically performed via Particle Image Velocimetry (PIV) with fluorescent polystyrene microspheres or Computational Fluid Dynamics (CFD), while local cell distribution is monitored through expensive sensors or direct sampling. However, PIV and CFD analysis lack of cell behaviour representativity, while direct sampling give average and local information and may impact cell culture conditions. In this study a novel non-invasive method, focusing on the optical investigation of suspended fluorescent nanoparticles (NPs) -labelled Chinese hamster ovary (CHO) cells distribution within SUSPENCE® bioreactor through PIV image processing, is presented. Our investigation showcases the favourable effect of an innovative NPs internalisation approach in terms of cellular uptake efficiency and fluorescence brightness. Moreover, NPs-labelled CHO cells (NP-CHO) PIV image processing and analysis robustness is validated by cell sampling and sample processing. Furthermore, the turbulent kinetic energy distribution to gain insight of the impact of hydrodynamic conditions on cell culture is evaluated. © 2022 Copyright for this paper by its authors.