3T3 Neutral Red Uptake cytotoxicity assay

Topic: Acute toxicity, Basal Cytotoxicity

Test Method Number:
TM2007-03 (EU)
Short Name of TM:
Year received:
Responsible Organisation:
General Comments:
The validation study assessed the capacity of the 3T3 NRU cytotoxicity assay to identify specifically non-classified chemicals on the basis of the 2000 mg/kg b.w. threshold introduced by the EU CLP Regulation.

Method Description

The 3T3 NRU cytotoxicity assay employs the BALB/c3T3 mouse fibroblast cell line and is based on the ability of viable cells to incorporate and bind the dye Neutral Red (NR), measured spectrophotometrically . The basic concept of basal cytotoxicity assays is that chemicals exert their toxic effects by disrupting structures and functions universal to all cells.
The assay used the test method protocol and the IC50-LD50 regressions of the NRU cytotoxicity method, validated in the NICEATM/ECVAM validation study (see TM2003-01) and was proposed to identify chemicals that would not require classification for acute oral toxicity according to the EU CLP Regulation (i.e. LD50 > 2 000 mg/kg). It was envisaged that use of the 3T3 NRU test method to identify unclassified substances in a testing strategy, to support a read-across, or as part of a weight-of-evidence approach could potentially reduce the amount of animal testing for acute oral toxicity.

Track Approval Status

    Regulatory acceptance/Standards
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