EURL ECVAM Recommendation on the 3T3 Neutral Red Uptake Cytotoxicity Assay for Acute Oral Toxicity Testing

Acute oral toxicity is currently being assessed by a suite of refinement test methods based on the traditional LD50 lethality test and is, besides skin sensitisation, the only remaining animal test required under REACH Annex VII. In view of assessing the use of alternatives for this health endpoint, EURL ECVAM conducted a study on the 3T3 Neutral Red Uptake cytotoxicity test method addressing the method's capacity to support specifically the identification substances not requiring classification as acute toxicants. Following independent scientific peer review of this study by EURL ECVAM's scientific advisory committee (ESAC) and having considered input from regulators, stakeholders, international partners and the general public, EURL ECVAM concludes that the 3T3 NRU test method may prove a valuable component of a WoE or ITS approach for supporting hazard identification and safety assessment in agreement with the EU CLP Regulation and international regulatory schemes implementing the upper threshold of UN GHS Category 4 as the cut-off for non-classification of substances. In particular, data from the 3T3 NRU assay may constitute an information source within a WoE approach under the provisions of the REACH regulation (Annex XI, 1.2) potentially supporting conclusions on absence of acute oral toxicity of industrial chemicals.JRC.I.5-Systems Toxicolog