The invention relates to a two-step temperature reduction method for eliminating low-temperature cracking by being preserved by a biological tissue vitrification method, belonging to the technical field of biomedical technology. The two-step temperature reduction method means that temperature reduction is firstly carried out in cold nitrogen and then transferred into liquid nitrogen. By adopting the two-step temperature reduction technology, the temperature reduction rate of the samples can be properly reduced, the heat stress in the samples can be weakened, and the low-temperature cracking is further expected eliminated; meanwhile, the vitrification of the samples can be still ensured. When two-step temperature reduction is implemented, the following operatioen is followed: in the first step of temperature reduction, the samples are placed in nitrogedn within the temperature range of -196 to -135 DEG C, and the temperature reduction rate is 10-30 DEG C/min within the temperature range of -80 to 4 DEG C, thereby ensuring the freeze-stored samples to realize vitrification in the process of temperature reduction; when the temperature is reduced within the range of -196 to -100 DEG C, the freeze-stored samples are immerged into the liquid nitrogen, and the temperature reduction rate is 0-165 DEG C/min within, thus preventing the main body part of the solution from cracking. The devices used for realizing the two-step temperature reduction are simple and comprise a Dewar bottle or similar equipment, and a temperature measuring device.