Abstract: As the primary immune cells, hemocytes are produced throughout the life cycle of crustacean. However, our understanding of crustacean hematopoiesis is limited. Recently, we found that the muscle extract of Cherax quadricarinatus can induce the differentiation of hematopoietic tissue (HPT) cell into granular cells in vitro. The exact factor that induces GC differentiation remains unknown. In this study, the basic properties of the unknown hemocytes differentiation factor were analyzed. We prepared muscle extracts from C. quadricarinatus, Eriocheir sinensis, Locusta migratoria manilensis, Epinephelus goreensis, Litopenaeus vannamei and Reishia clavigera. Their ability to induce HPT cells differentiation was tested. Results showed that all the muscle extracts could induce the differentiation of HPT cells along the GCs lineage, indicating that the hemocyte differentiation factor exists in all tested animals. Further, we analyzed the thermal stability, solubility, electrification, resistance to protease and molecular weight of the hemocytes differentiation factor in the muscle extract of C. quadricarinatus. We found that the target factor was a small molecule (< 1 kDa), which was methanol-soluble, electrically neutral, thermal stable and protease-resistant. Therefore, the target factor is a widely existing small molecule. Thus, results of this study should facilitate the purification and identification of this factor.