Mouse N2a Neuroblastoma Assay: Uncertainties and Comparison with Alternative Cell-Based Assays for Ciguatoxin...

Area of Science:

• Food safety
• Marine toxicology
• Cell biology

Background:

• Ciguatera fish poisoning (CF) is a growing concern due to ciguatoxins (CTXs).
• Current N2a cell-based cytotoxicity assays for CTX detection lack standardization and have limitations.
• N2a cells show low sodium channel availability and are susceptible to damage from ouabain (O) and veratridine (V).

Purpose of the Study:

• To evaluate alternative cell lines for reliable and standardized ciguatoxin detection.
• To identify a cell line sensitive to CTXs and suitable for cytotoxicity assays.
• To improve food safety by developing a faster method for ciguatoxin analysis.

Main Methods:

• Evaluation of N2a cells and alternative cell lines (human neuroblastoma, HEK293 Nav).
• Assessment of cell viability using classical cytotoxicity assays (MTT) and methods without medium removal (LDH, Alamar blue).
• Testing sensitivity to CTXs with and without ouabain (O) and veratridine (V) sensitization.

Main Results:

• N2a cells exhibited low sodium channel availability and significant damage from O/V.
• Human neuroblastoma cells were unsuitable due to high O/V sensitivity and low sodium channel expression.
• HEK293 Nav cells, expressing sodium channels, were sensitive to CTXs without O/V sensitization.
• MTT assay with O/V sensitization detected CTX3C at 0.0001 nM in HEK293 Nav cells.

Conclusions:

• The HEK293 Nav cell line is a viable alternative for ciguatoxin detection.
• This cell line offers improved sensitivity and reliability compared to traditional methods.
• This advancement contributes to better food safety and monitoring of ciguatoxins.