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Updated: Jun 10, 2026

Fluorescent Leakage Assay to Investigate Membrane Destabilization by Cell-Penetrating Peptide
Published on: December 19, 2020
Development of a fixed-dose approach for the fluorescein leakage test
R H Clothier1, S J Morgan, K A Atkinson
1FRAME Alternatives Laboratory, Department of Human Morphology, University of Nottingham Medical School, Nottingham NG7 7UH, UK.
Abstract:
To answer the question, "How toxic is this material?", rather than the question more usually asked in in vitro toxicology, "At what concentration does this material produce a given degree of toxicity?", a fixed-dose approach has been developed with the FRAME fluorescein leakage (FL) test. Recovery from the initial cytotoxic effect was also noted sometimes, after the cells had been maintained in fresh medium for a further 72 hr. Chemicals rated R41 or R36 in vivo gave a level of FL above 20% immediately, and 66% or more 72 hr later (i.e. there was further deterioration, rather than recovery). Acetaldehyde, when duplicated and tested either in Hanks' balanced salt solution or oil, showed greater toxicity in the oil because of its high volatility. The chemicals classified as no-label gave FL values below 12% immediately after exposure, except Brij-35, which showed no further deterioration during the 72-hr period in fresh medium.
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