The toxicity of CdTe QDs with nearly identical maximum emission wavelength but modified with four different ligands (MPA, NAC, GSH and dBSA) to HEK293 and HeLa cells were investigated using flow cytometry, spectroscopic and microscopic methods. The results showed that the cytotoxicity of QDs increased in a dose- and time-dependent manner. No appreciable fraction of cells with sub-G1 DNA content, the loss of membrane integrity, and the swelling of nuclei clearly indicated that CdTe QDs could lead to necrotic cell death in HEK293 cells. JC-1 staining and TEM images confirmed that QDs induced MPT, which resulted in mitochondrial swelling, collapse of the membrane potential. MPT is an important step in QDs-induced necrosis. Moreover, QDs induced MPT through the elevation of ROS. The fluorimetric assay and theoretical analysis demonstrated ROS production has been associated with the internalization of QDs with cells. Due to large surface/volume ratios of QDs, when QDs added in the culture medium, serum proteins in the culture medium will be adsorbed on the surface of QDs. This adsorption of serum protein will change the surface properties and size, and then mediate the cellular uptake of QDs via the clathrin-mediated endocytic pathway. After entering into cells, the translocation of QDs in cells is usually via endosomal or lysosomal vesicles. The rapid degradation of QDs in lysosome and the lysosomal destabilization induce cell necrosis. This study provides a basis for understanding the cytotoxicity mechanism of CdTe QDs, and valuable information for safe use of QDs in the future.