【Objective】In previous studies, OsHSP20, a gene encoding small heat shock protein, was cloned from Oryza sativa. Here its polyclonal antibodies were prepared and analyzed for their properties of immunological reactions and range of application. 【Method】 Synthetic peptide and recombinant protein of OsHSP20 were used to immunize the rabbits for preparing the polyclonal antibodies against the protein, respectively. Antigen-coated plate (ACP)- and dot-enzyme linked immunosorbent assays (ELISA) were used for comparing the titer and sensitivity of antibodies. Western blotting was used to determine the antibody specificity and the range of application. 【Result】In Western blotting assays, both antibodies were strongly reacted with a band of protein with an expected size of 37 kD, suggesting that they were specific against the OsHSP20. ACP- and dot-ELISA assays showed that the polyclonal antibody prepared from the recombinant protein had a higher titer and sensitivity than that of peptide. Further assays showed that the polyclonal antibody could be used for detection of the SHSP homologs of plants. 【Conclusion】The polyclonal antibodies were successfully prepared with high specificity, titer, and sensitivity and could be widely used to analyze the expression of HSP20 protein in a variety of monocotylous and dicotyledonous plants, which could contribute to the functional characterization of OsHSP20 and its homologs in different plants.