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In this study, whether Giemsa staining solution can accurately determine bacterial contamination of liquid spawn for Flammulina velutipes in a short period of time was investigated. Giemsa solution staining cells of blood, bone marrow, lymph node, malaria parasites, rickettsia et al. was prepared by dissolving basic methylene azul and methylene blue, and acidic eosine in methyl alcohol-glycerine. Supernatant samples of Flammulina velutipes liquid spawn cultured under explosive aeration were placed on a slide, mixed with Gimesa solution and examined with optical microscope after staining. In 40 to 60 seconds bacterial cells were distinguishable from soybean meal residual and hyphal cell fragments. Thus we conclude that microscopy using Gimesa staining solution is a quick, simple and accurate method for the mushroom growers to effectively use to detect bacterial contamination of the liquid spawn.