Abstract

Ethical concerns regarding the mouse bioassay (AOAC 959.08), the toxicological assay for the detection of paralytic shellfish toxins (PST), have prompted the search for new toxicological methodologies in order to eliminate the use of animals for monitoring. One of the promising approaches is the use of in vitro toxicological methods such as the cell-based assay using mouse neuroblastoma Neuro-2a cells (CBA). However, one of the problems of this assay is the effect produced by the shellfish meat (matrix), so it is necessary to evaluate the effect of the extraction method before adapting the CBA to natural samples. The aim of this study was to evaluate the use of CBA as a screening tool for the detection of PST in mussel contaminated samples. The first step was to evaluate the matrix effect in the Neuro-2A cells, comparing three extraction protocols (AOAC 2005.06, AOAC 2011.02 and liquid-liquid extraction method). Then, using the AOAC 2005.06 extraction protocol, which presented less matrix effect, a prevalidation of the CBA was performed. For this purpose, the repeatability and reproducibility of the assay was measured using reference material. Both parameters fulfill the criteria of acceptability at matrix concentrations between 1.5 and 25 mg meat/mL. Finally, the discrimination capacity of the CBA using the cutoff value obtained from fortified samples was evaluated. A total of 13 natural PST-contaminated samples were evaluated, and 11 were detected by the CBA. These results suggest the usefulness of CBA as a screening method, although it is necessary to complete the validation process.