Polymeric nanoparticles (NPs) are promising tools for immunomodulation and drug delivery across various disease contexts. However, despite their pre-clinical success, clinical translation remains limited. This is often attributed to insufficient NP characterization and inconsistent experimental reporting, leading to variability in literature and reproducibility issues. Although extensive research has been conducted, the factors governing nano-bio interactions are still not fully understood. One factor strongly associated with impaired NP efficacy is how endogenous biomolecules interact with their surfaces. Upon exposure to biological environments, plasma-resident biomolecules rapidly coat NPs, forming a biomolecular corona that varies between individuals and is influenced by NP formulation parameters. This dissertation explores key aspects of the NP development process that influence biological interactions. First, we demonstrate that altering formulation parameters, such as purification and synthesis methods, can produce NPs with similar physicochemical properties but markedly different cellular interactions. Notably, tangential flow filtration (TFF) purification and tuning flow rate ratios (FRRs) during microfluidic synthesis significantly modulates NP corona composition and influences organ distribution despite similar NP characteristics. Next, we investigate how inflammatory disease progression impacts NP corona composition and its effects on innate immune activation. Our findings reveal that disease-associated changes in the corona affect cell interactions, modulate co-stimulatory and co-inhibitory molecule expression, and alter cytokine release through TLR4/MyD88/NF-κB signaling pathways. Finally, we evaluate the use of pre-formed lactoferrin coronas to enhance antimicrobial activity and improve pathogen clearance of previously established anti-inflammatory NPs. Together, these findings highlight the formulation-dependent and personalized nature of corona formation, emphasizing how minor variations in the NP development process can lead to significant differences in biological outcomes and immunogenicity.