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Alternatives to Laboratory Animals - ATLA

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The Neutral Red Release Assay: A Review


Valérie Zuang

The neutral red release (NRR) assay is a cytotoxicity test that can be used to measure the immediate toxic effects of test substances on the cell membrane, resulting in the leaking of intracellular contents. The assay has already been used for several years to evaluate the cytotoxicities of various kinds of products, such as cosmetics, pharmaceuticals, industrial chemicals and household products. It has undergone in-house validation by many companies, and has been found to be particularly useful for identifying substances that are potentially capable of causing adverse reactions on coming into brief contact with the eye or the skin at relatively high concentrations, such as might occur in an adventitious splash into the eye or onto the skin, followed by a quick rinse. Because of the relatively long existence of the NRR assay, its practicality and its proven usefulness for particular purposes, ECVAM decided to review the status of the method, in order to decide whether prevalidation and formal validation studies on the test might be profitable. The review of the status of the test was carried out by performing a comprehensive review of the literature, and by conducting a survey involving companies and institutes with experience in using the test. Both the review and the survey revealed that the assay could provide extremely valuable information when it was used for particular purposes, such as for the evaluation and comparison of immediate toxic effects on the eye or the skin caused by certain products or chemicals such as surfactants. Most of those who responded in the survey favoured a prevalidation/validation study.