A nanoparticle's physical and chemical properties at the time of cell contact will determine the ensuing cellular response. Aggregation and the formation of a protein corona in the extracellular environment will alter nanoparticle size, shape, and surface properties, giving it a "biological identity" that is distinct from its initial "synthetic identity". The biological identity of a nanoparticle depends on the composition of the surrounding biological environment and determines subsequent cellular interactions. When studying nanoparticle-cell interactions, previous studies have ignored the dynamic composition of the extracellular environment as cells deplete and secrete biomolecules in a process known as "conditioning". Here, we show that cell conditioning induces gold nanoparticle aggregation and changes the protein corona composition in a manner that depends on nanoparticle diameter, surface chemistry, and cell phenotype. The evolution of the biological identity in conditioned media enhances the cell membrane affinity, uptake, and retention of nanoparticles. These results show that dynamic extracellular environments can alter nanoparticle-cell interactions by modulating the biological identity. The effect of the dynamic nature of biological environments on the biological identity of nanoparticles must be considered to fully understand nano-bio interactions and prevent data misinterpretation.