we set protein corona as the research object in this study, and focused on exploring the biological effects of intracellular protein corona (IPC) that formed on particle surface after transcytosis of nanomedicines. Considering the multi-functionalization characteristic, gold nanoparticles (AuNPs) were fabricated here as the model of nanomedicines. We established a dual-filtration proteomics strategy based on mass spectrometry (MS) to detect the composition and category of IPC after transcytosis. Based on the analysis on IPC proteins, we did achieve novel and significant findings.