The invention discloses a preservation method of a hilsa herring sperm. The preservation method comprises mixing an antifreeze agent, the hilsa herring sperm and a diluent at the volume ratio of 1:1:5, and preserving at the temperature of 5 DEG C, wherein the diluent comprises the ingredients in percent by volume: 50% of sodium chloride injection, 15-20% of pure water, 7-14% of potassium chloride,5-10% of calcium chloride, 5-10% of sodium bicarbonate, 1-2% of chlorogenic acid, 1-3% of glycerinum, 3-5% of glucose, 8-15% of lecithin, 1-3% of sodium citrate, 1-2% of streptomycin, 1-3% of poly-sodium glutamate, 1-3% of bifidobacterium, and the balance of lactic acid bacteria by volume; and the antifreeze agent comprises the ingredients in percent by volume: 30% of dimethyl sulfoxide, 20% of glycerinum, 30% of ethylene glycol, and 20% of dimethylacetamide. The preservation method effectively inhibits the growth and propagation of harmful microbes, reduces the metabolic rate of the sperm, effectively keeps the preservation quality of the hilsa herring sperm, and saves the production cost, and operation steps are convenient.