Bioconjugated nanoparticles are commonly used for targeting cellular/subcellular components, and labeling performance is known to depend on multivalency, i.e., the number of attached biomolecule per particle. However, these multivalency effects are largely unexplored. Here, we show that multivalency of nanoparticle-bound riboflavin controls the cellular interaction, cellular entry/exit mechanism, and subcellular trafficking property. We have synthesized riboflavin-functionalized quantum dot (QD) of 15-25 nm hydrodynamic size with average riboflavin multivalencies of 15, 30, and 70 [designated as QD(RF)15, QD(RF)30, and QD(RF)70, respectively] and investigated their uptake mechanism in riboflavin receptor overexpressed KB cells. We found that increased multivalency from 15 to 70 increases the cellular interaction with QD, shifts the cell uptake mechanism from caveolae-clathrin to exclusive clathrin-mediated endocytosis, and enhances lysosomal trafficking. This work demonstrates the importance of multivalency of bioconjugated molecule at the nanoparticle surface toward biolabeling performance and should be optimized for best performance of designed nanobioconjugate.