A method for the determination of tetrodotoxin (TTX) in aquatic foods using hydrophilic interaction liquid chromatography-tandem mass spectrometry (LC-MS/MS) was been established.Methods The samples were extracted by 1% acetic acid-methanol and diluted with phosphate buffer, followed by the purification and concentration process by immunoaffinity columns (IACs). The separation of TTX was carried out on a column of TSK-gel Amide-80, and detected by positive electrospray ionization tandem mass spectrometry in the SRM mode. The external solvent standard calibration curves were used for quantitative analysis. Results The calibration curves were linear in the range of 1-1 000 μg/L for TTX, with the detection limit of 1 μg/kg and the quantification limit of 3 μg/kg. The average recoveries were between 73.6% and 95.2% with relative standard deviations (RSD) ranged from 5.37% to 10.7%.Conclusion The matrix effect was substantially eliminated and the stability of retention time were improved obviously. The proposed method could be used to quantify and identify TTX in fish products with excellent sensitivity, accuracy and reproducibility.