The invention belongs to the chemical field, and provides a preparation method for monodispersed and low-biotoxicity gold nanorods, and use for detection of allergen. The preparation method comprises the steps of (1) adding cetyltrimethylammonium bromide (CTMBA) in a beaker, adding HAuCl4 and NaBH4 and stirring; (2) adding a growth solution and an anionic surfactant into the beaker, adding HAuCl4 and ascorbic acid with stirring, adding gold seeds prepared by the step (1), heating in a water bath, cooling, and filtering precipitated CTMAB; (3) taking the gold nanorods prepared by the step (2), adding a PBS solution and a probe DNA and putting in a refrigerator; and (4) taking a certain amount of the modified gold nanorods prepared by the step (3), adding a target DNA, measuring synchronous fluorescence and calculating the content of allergen according to a regression curve. According to the preparation method, the anionic surfactant is used as an additive, so that the usage amount of CTMAB is reduced. The prepared gold nanorods have good monodispersity and low biotoxicity. By using the gold nanorods in the detection of allergen, a detection limit and sensitivity of the method is obviously higher than that of a conventional technology.