The aim of this work was to develop a liquid chromatography-tandem mass spectrometry method for the determination of milk allergen and egg allergen in food products. Signature peptides GGLEPINFQTAADQAR, VGINYWLAHK, VLVLDTDYK, FFVAPFPEVFGK, and NAVPITPTLNR were confirmed and synthesized as the quantitative peptide of ovalbumin, α-lactalbumin, β-lactoglobulin, αS1-casein and αS2-casein, the relative isotope-labeled internal standards were used in the quantitative analysis. Linear range was in the range of 0.5–5 000.0 nmol/L for egg and milk allergen in bread, cake, cookie, rice crust and wheat flour samples with free from egg and milk, the limits of detection of milk allergens and egg allergen were in the range between 0.94 mg/100 g and 56.71 mg/100 g, limits of quantification of milk allergens and egg allergen were in the range between 2.36 mg/100 g and 141.78 mg/100 g. The recoveries ranged from 76.7% to 122.8%, the relative standard deviations were in the range of 1.60%–15.60%. The developed method has been successfully used for the detection of egg and milk allergen in various food samples.