Applications of the Neutral Red Cytotoxicity Assay to Risk Assessment of Aquatic Contaminants: An Overview

Abstract

The neutral red (NR) in vitro cell viability assay is a cytotoxicity test, initially developed for use with mammalian cells to evaluate the acute toxicities of chemicals. It has been adapted for aquatic ecotoxicity tests by the use of cultured fish cells as the bioindicator system. This assay is based on the binding of neutral red, a weakly cationic, supravital dye, to the lysosomal matrix of viable cells after their incubation with toxic agents. Spectrophoto-metric quantitation of the extracted dye at 540 nm with a scanning microtiter well reader was shown to be linear with the number of surviving, undamaged, viable cells. This assay with fish cells as the targets has been applied to five areas of ecotoxicity testing and risk assessment: (1) the ranking of the test agents according to their potencies; (2) the study of metabolism-mediated cytotoxicity; (3) the analysis of structure-activity relationships for series of related chemicals; (4) the determination of chemical toxicity as a function of temperature; and (5) the evaluation of chemical interactions as they relate to toxicity.

A comparison of BGM and LLC-PK1 cells for the evaluation of nephrotoxicity

“…The test with NR was performed based on the studies of Boreunfreund and Puerner (1984) and Babich and Borenfreund (1993). The NR uptake assay provides a quantitative estimation of the number of viable cells in a culture.…”