In vitro cytotoxicity assays: comparison of LDH, neutral red, MTT and protein assay in hepatoma cell lines following exposure to cadmium chloride

Toxicol Lett. 2006 Jan 5;160(2):171-7. doi: 10.1016/j.toxlet.2005.07.001. Epub 2005 Aug 18.


The aim of this study was to compare four in vitro cytotoxicity assays and determine their ability to detect early cytotoxic events. Two hepatoma cell lines, namely HTC and HepG2 cells, were exposed to cadmium chloride (0-300 microM) for 3, 5 and 8 h. Following exposure to the toxic metal cytotoxicity was determined with the lactate dehydrogenase leakage assay (LDH), a protein assay, the neutral red assay and the methyl tetrazolium (MTT) assay. In HTC cells no toxicity was observed for any incubation period when the LDH leakage, the MTT and the protein assay were employed whereas the neutral red assay revealed early cytotoxicity starting after incubation of HTC cells with CdCl(2) for 3 h. In the case of HepG2 cells the MTT assay reveals cytotoxicity due to CdCl(2) exposure after 3 h whereas no such effect is seen with the other three assays. Following 5 h exposure of HepG2 cells to CdCl(2), toxicity is observed with the MTT assay at lower concentrations compared to the ones required for detection of toxicity with the LDH leakage and the neutral red assay. In conclusion different sensitivity was observed for each assay with the neutral red and the MTT assay being the most sensitive in detecting cytotoxic events compared to the LDH leakage and the protein assay.

Publication types

  • Comparative Study

MeSH terms

  • Animals
  • Cadmium Chloride / toxicity*
  • Carcinoma, Hepatocellular
  • Cell Line, Tumor
  • Cell Survival / drug effects*
  • Coloring Agents
  • Humans
  • L-Lactate Dehydrogenase / analysis
  • L-Lactate Dehydrogenase / metabolism
  • Neutral Red
  • Proteins / analysis
  • Proteins / metabolism
  • Rats
  • Tetrazolium Salts
  • Thiazoles
  • Toxicity Tests*


  • Coloring Agents
  • Proteins
  • Tetrazolium Salts
  • Thiazoles
  • Neutral Red
  • L-Lactate Dehydrogenase
  • thiazolyl blue
  • Cadmium Chloride