This paper presents a simple spectroscopic method for differentiating cellular uptakes of two different types of Au nanostructures (i.e., nanospheres and nanorods) when they are supplied as a mixture. Because Au nanospheres and nanorods have distinctive surface plasmon resonance peaks, we can simultaneously measure their concentrations in a culture mediumbyUV-Vis spectroscopy. For the uptake of Au nanostructures by SK-BR-3 breast cancer cells, we found that the surface functional group had a stronger influence than the shape. We also found that the cellular uptake of one type of Au nanostructure could be affected bythe presence of another type of nanostructure in the medium. This study not only suggests a simple way to compare the cellular uptakes of more than one type of Au nanostructures without incubating them separately but also allows one to investigate the influence of one type of Au nanostructures on the uptake of another type.