Gao Li-juan, Yang Wei-dong, Liu Jie-sheng
Department of Biotechnology, Jinan University, Guangzhou 510632, China.
Guang Pu Xue Yu Guang Pu Fen Xi. 2009 Apr;29(4):1032-5.
We developed a method to screen paralytic shellfish poisoning (PSP) toxins based on their functional activity. The assay was a fluorimetric assay by detecting changes in the membrane potential of transitional cell carcinoma of the bladder cells T24 and involved several steps: stain of T24 cells with fluorescent dye bis-oxonol, cell depolarization with veratridine, and inhibition of depolarization with PSP toxins GTX2, 3 or shellfish samples containing PSP toxins. Toxic potency of the samples was evaluated by measuring toxin-induced changes in membrane potential. Within 2-100 nmol x L(-1) of GTX2, 3, veratridine-induced depolarization was shown to be inhibited by GTX2, 3 in a dose-dependent manner. There was a linear correlation between the percentage of inhibition and toxin concentration. The PSP toxin value in shellfish obtained by this fluorescence assay was in concordance with that by the mouse bioassay, and with higher sensitivity. In conclusion, the fluorescent dye method based on changes in membrane potential was a rapid, specific, and reliable method for detecting paralytic shellfish poisoning toxins in shellfish.
我们开发了一种基于麻痹性贝类毒素（PSP）功能活性来筛选该毒素的方法。该检测方法是一种荧光检测法，通过检测膀胱移行细胞癌T24细胞的膜电位变化来进行，涉及多个步骤：用荧光染料双苯甲酰羟肟对T24细胞进行染色，用藜芦定使细胞去极化，并用PSP毒素GTX2、3或含有PSP毒素的贝类样本抑制去极化。通过测量毒素诱导的膜电位变化来评估样本的毒性强度。在GTX2、3浓度为2 - 100 nmol x L(-1)范围内，藜芦定诱导的去极化被GTX2、3以剂量依赖的方式抑制。抑制百分比与毒素浓度之间存在线性相关性。通过这种荧光检测法获得的贝类中PSP毒素值与小鼠生物检测法的结果一致，且灵敏度更高。总之，基于膜电位变化的荧光染料法是一种快速、特异且可靠的检测贝类中麻痹性贝类毒素的方法。
