This study investigated the cellular uptake of fluorescein isothiocyanate-labelled mesoporous silica nanoparticles (FITC-MSNs), amine-functionalised FITC-MSNs (AP-FITC-MSNs) and their gallic acid (GA)-loaded counterparts. Mesoporous silica nanoparticles were labelled with fluorescein isothiocyanate, functionalised by 3-aminopropyltriethoxysilane (APTES) (AP-FITC-MSNs) and then loaded by GA. All nanoparticles were characterised by transmission electron microscopy (TEM), Fourier transform infrared spectroscopy, and X-ray diffraction. The cytotoxicity of different concentrations of dyed nanoparticles was investigated using (3-(4,5-trihydroxybenzoic acid, dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and flow cytometry. TEM images showed that the average particle sizes of FITC-MSNs and AP-FITC-MSNs were about 100 and 110 nm, respectively. These nanoparticles were internalised by Caco-2 cells, accumulated and dispersed into the cytoplasm, nucleus, and subcellular organelles. Nanoparticles containing GA clearly decreased the viability of cells. FITC-MSNs showed no toxicity on Caco-2 cells at concentrations of ≤50 µg/ml. Functionalisation of FITC-MSNs using APTES decreased toxicity effects on the cells. It was found that FITC-MSNs can be applied at low concentrations as a marker in the cells. In addition, AP-FITC-MSNs showed better biocompatibility with Caco-2 cells than FITC-MSNs, because of their positive surface charges.