The impact of nanomaterials such as carbon nanotubes on biological matter is a topic of increasing interest and concern and requires a multifaceted approach to be resolved. A modiﬁ ed cytotoxic (lactate dehydrogenase (LDH)) assay is developed in an attempt to offer a valid and reliable methodology for screening carbon nanotube toxicity in vitro. Two of the most widely used types of surface-modiﬁ ed multiwalled carbon nanotubes (MWNTs) are tested: ammonium-functionalized MWNTs (MWNT-NH 3+) and Pluronic F127 coated MWNTs (MWNT:F127). Chemically functionalized MWNTs show signiﬁ cantly greater cellular uptake into lung epithelial A549 cells compared to the non-covalently Pluronic F127-coated MWNTs. In spite of this, MWNT:F127 exhibit enhanced cytotoxicity according to the modiﬁ ed LDH assay. The validity of the modiﬁ ed LDH assay is further validated by direct comparison with other less reliable or accurate cytotoxicity assays. These ﬁ ndings indicate the reliability of the modiﬁ ed LDH assay as a screening tool to assess carbon nanotube cytotoxicity and illustrate that high levels of carbon nanotube cellular internalization do not necessarily lead to adverse responses.