The invention discloses a preparation method for a fugu rubripes uronema marinum disease vaccine. The preparation method comprises the following steps that: picking up a single uronema marinum to be inoculated to an L-15 complete medium to be subjected to pure culture; then, carrying out enlargement cultivation; adding 100 mu l/L of formalin to kill larvae; carrying out centrifugation for 10 min at 4000rpm by a low-speed centrifugal machine to collect sediments; adding into the L-15 complete medium to prepare inactivation larva liquid of which the density is 2 million/ml; collecting in a centrifuge tube of 1.5ml, and preserving at the temperature of -80 DEG C. Before immunization is carried out, dead larvae are unfrozen on ice, are broken by electromagnetic wave and are mixed with equivalent Freund's adjuvant or Freund's incomplete adjuvant, and the mixed liquid is oscillated to emulsion to prepare the fugu rubripes uronema marinum disease vaccine. Since an intraperitoneal injection way is adopted, the immunity of the fugu rubripes for the uronema marinum disease can be improved.