Enhanced understanding of bio-nano interaction requires recognition of hidden factors such as protein corona, a layer of adsorbed protein around nano-systems. This study compares the biological identity and fingerprint profile of adsorbed proteins on PLGA-based nanoparticles through nano-liquid chromatography-tandem mass spectrometry. The total proteins identified in the corona of nanoparticles (NPs) with different in size, charge and compositions were classified based on molecular mass, isoelectric point and protein function. A higher abundance of complement proteins was observed in modified NPs with an increased size, while NPs with a positive surface charge exhibited the minimum adsorption for immunoglobulin proteins. A correlation of dysopsonin/opsonin ratio was found with cellular uptake of NPs exposed to two positive and negative Fc receptor cell lines. Although the higher abundance of dysopsonins such as apolipoproteins may cover the active sites of opsonins causing a lower uptake, the correlation of adsorbed dysopsonin/opsonin proteins on the NPs surface has an opposite trend with the intensity of cell uptake. Despite the reduced uptake of corona-coated NPs in comparison with pristine NPs, the dysopsonin/opsonin ratio controlled by the physicochemistry properties of NPs could potentially be used to tune up the cellular delivery of polymeric NPs.