In the present study, we compared cytotoxicity and cell uptake of silica nanoparticles with four different surface coatings generated through layer-by-layer self-assembly. Rabbit mesenchymal stem cells (rMSCs) were labeled with silica nanoparticles of different coatings including poly(ethyleneimine) (PEI), poly(allylamine hydrochloride) (PAH), poly(anetholesulfonic acid, sodium salt) (PAS), and dextran sulfate. The MTT [3-(4, 5-dimethylthiazol-2)-2, 5-diphenyl-2H-tetrazolium bromide] test was performed to quantify the cell biocompatibility. The cellular uptake of those silica nanoparticles was determined by flow cytometry and confocal laser scanning microscopy. The results showed that all examined silica nanoparticles were stable in aqueous phase with high monodispersity. Labeled rMSCs are unaffected in their viability, apoptosis, and differentiation capacities. The silica nanoparticle-coated synthetic polycations such as PEI or PAH have higher cell internalization than negatively charged polyelectrolytes. The ability to control cell uptake of different particles may have applications in cell labeling, cell separation, and other biomedical applications.