Tags

Type your tag names separated by a space and hit enter

Revisiting the Neuroblastoma Cell-Based Assay (CBA-N2a) for the Improved Detection of Marine Toxins Active on Voltage Gated Sodium Channels (VGSCs).
Toxins (Basel). 2020 04 27; 12(5)T

Abstract

The neuroblastoma cell-based assay (CBA-N2a) is widely used for the detection of marine biotoxins in seafood products, yet a consensus protocol is still lacking. In this study, six key parameters of CBA-N2a were revisited: cell seeding densities, cell layer viability after 26 h growth, MTT incubation time, Ouabain and Veratridine treatment and solvent and matrix effects. A step-by-step protocol was defined identifying five viability controls for the validation of CBA-N2a results. Specific detection of two voltage gated sodium channel activators, pacific ciguatoxin (P-CTX3C) and brevetoxin (PbTx3) and two inhibitors, saxitoxin (STX) and decarbamoylsaxitoxin (dc-STX) was achieved, with EC50 values of 1.7 ± 0.35 pg/mL, 5.8 ± 0.9 ng/mL, 3 ± 0.5 ng/mL and 15.8 ± 3 ng/mL, respectively. When applied to the detection of ciguatoxin (CTX)-like toxicity in fish samples, limit of detection (LOD) and limit of quantification (LOQ) values were 0.031 ± 0.008 and 0.064 ± 0.016 ng P-CTX3C eq/g of flesh, respectively. Intra and inter-assays comparisons of viability controls, LOD, LOQ and toxicity in fish samples gave coefficients of variation (CVs) ranging from 3% to 29%. This improved test adaptable to either high throughput screening or composite toxicity estimation is a useful starting point for a standardization of the CBA-N2a in the field of marine toxin detection.

Authors+Show Affiliations

Institut Louis Malardé (ILM), Laboratory of Marine Biotoxins-UMR 241-EIO, 98713 Papeete-Tahiti, French Polynesia.Institut Louis Malardé (ILM), Laboratory of Marine Biotoxins-UMR 241-EIO, 98713 Papeete-Tahiti, French Polynesia.Institut Louis Malardé (ILM), Laboratory of Marine Biotoxins-UMR 241-EIO, 98713 Papeete-Tahiti, French Polynesia.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Validation Study

Language

eng

PubMed ID

32349302

Citation

Viallon, Jérôme, et al. "Revisiting the Neuroblastoma Cell-Based Assay (CBA-N2a) for the Improved Detection of Marine Toxins Active On Voltage Gated Sodium Channels (VGSCs)." Toxins, vol. 12, no. 5, 2020.
Viallon J, Chinain M, Darius HT. Revisiting the Neuroblastoma Cell-Based Assay (CBA-N2a) for the Improved Detection of Marine Toxins Active on Voltage Gated Sodium Channels (VGSCs). Toxins (Basel). 2020;12(5).
Viallon, J., Chinain, M., & Darius, H. T. (2020). Revisiting the Neuroblastoma Cell-Based Assay (CBA-N2a) for the Improved Detection of Marine Toxins Active on Voltage Gated Sodium Channels (VGSCs). Toxins, 12(5). https://doi.org/10.3390/toxins12050281
Viallon J, Chinain M, Darius HT. Revisiting the Neuroblastoma Cell-Based Assay (CBA-N2a) for the Improved Detection of Marine Toxins Active On Voltage Gated Sodium Channels (VGSCs). Toxins (Basel). 2020 04 27;12(5) PubMed PMID: 32349302.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Revisiting the Neuroblastoma Cell-Based Assay (CBA-N2a) for the Improved Detection of Marine Toxins Active on Voltage Gated Sodium Channels (VGSCs). AU - Viallon,Jérôme, AU - Chinain,Mireille, AU - Darius,Hélène Taiana, Y1 - 2020/04/27/ PY - 2020/04/03/received PY - 2020/04/23/revised PY - 2020/04/24/accepted PY - 2020/5/1/entrez PY - 2020/5/1/pubmed PY - 2021/3/3/medline KW - CBA-N2a KW - absorbance data KW - biological sample KW - brevetoxins KW - ciguatoxins KW - matrix effects KW - saxitoxins KW - seafood safety KW - standardization JF - Toxins JO - Toxins (Basel) VL - 12 IS - 5 N2 - The neuroblastoma cell-based assay (CBA-N2a) is widely used for the detection of marine biotoxins in seafood products, yet a consensus protocol is still lacking. In this study, six key parameters of CBA-N2a were revisited: cell seeding densities, cell layer viability after 26 h growth, MTT incubation time, Ouabain and Veratridine treatment and solvent and matrix effects. A step-by-step protocol was defined identifying five viability controls for the validation of CBA-N2a results. Specific detection of two voltage gated sodium channel activators, pacific ciguatoxin (P-CTX3C) and brevetoxin (PbTx3) and two inhibitors, saxitoxin (STX) and decarbamoylsaxitoxin (dc-STX) was achieved, with EC50 values of 1.7 ± 0.35 pg/mL, 5.8 ± 0.9 ng/mL, 3 ± 0.5 ng/mL and 15.8 ± 3 ng/mL, respectively. When applied to the detection of ciguatoxin (CTX)-like toxicity in fish samples, limit of detection (LOD) and limit of quantification (LOQ) values were 0.031 ± 0.008 and 0.064 ± 0.016 ng P-CTX3C eq/g of flesh, respectively. Intra and inter-assays comparisons of viability controls, LOD, LOQ and toxicity in fish samples gave coefficients of variation (CVs) ranging from 3% to 29%. This improved test adaptable to either high throughput screening or composite toxicity estimation is a useful starting point for a standardization of the CBA-N2a in the field of marine toxin detection. SN - 2072-6651 UR - https://www.unboundmedicine.com/medline/citation/32349302/Revisiting_the_Neuroblastoma_Cell_Based_Assay__CBA_N2a__for_the_Improved_Detection_of_Marine_Toxins_Active_on_Voltage_Gated_Sodium_Channels__VGSCs__ L2 - https://www.mdpi.com/resolver?pii=toxins12050281 DB - PRIME DP - Unbound Medicine ER -