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目的:麻痹性贝类毒素的N-2a细胞检测方法的建立并确定可能的检测限值。方法:使用不同浓度毒素标准品结合藜芦定和乌本苷共同作用对数生长期的N-2a细胞,通过cck-8试剂盒检测细胞毒性,确立细胞检测方法的检出限及比较各毒素作用大小,并同步利用小鼠生物法进行验证,通过精确记录小鼠死亡时间,比较各毒素的毒作用大小。结果:麻痹性贝类毒素能明显降低藜芦定和乌本苷的细胞毒性,且具有剂量-反应关系,通过比较得出毒素各成分毒性大小为:neoSTX>STX>dcSTX>GTX1,4>GTX2,3>dcGTX2,3;小鼠生物法的毒作用平均死亡时间为:neoSTX组(6.5 min),GTX1,4组(8.0 min),STX组(9.0 min),dcSTX组(15 min);GTX2,3组和dcGTX2,3组未见动物死亡。结论:细胞检测与小鼠生物法具有较好的一致性,表明所建立的细胞检测法可行,且细胞毒性试验检测方法具有较高的灵敏度,最低检出限值可达到10-9mol/L剂量水平,其中浓度在10-6~10-8mol/L间具有较好的线性。
OBJECTIVE: To establish a method for the detection of paralytic shellfish toxin N-2a cells and to determine possible detection limits. Methods: N-2a cells in logarithmic growth phase were treated with verapamil and vinorelbine at different concentrations. Cytotoxicity was detected by cck-8 kit, and the detection limits of the cells were established. The size of the effect, and the simultaneous use of mouse biological method to verify, by accurately recording the time of death in mice to compare the toxicity of each toxin size. Results: Paralytic shellfish toxin could significantly reduce the cytotoxicity of veratridine and ouabain, and had a dose-response relationship. The toxicity of each component of toxin was neoSTX> STX> dcSTX> GTX1,4> GTX2 , 3> dcGTX2,3. The average lethal time of toxic effects in mice was: neoSTX group (6.5 min), GTX1,4 group (8.0 min), STX group (9.0 min), dcSTX group , 3 groups and dcGTX2, 3, no animals died. CONCLUSION: The cell test has good consistency with the biological test in mice, indicating that the established cell test is feasible and the cytotoxicity assay has high sensitivity with the lowest detection limit of 10-9 mol / L Level, in which the concentration of 10-6 ~ 10-8mol / L between the better linear.