In various radiation accidents, good estimation of absorbed dose in the victims is very important and influences the medical treatment decisions. Dicentric assay (metaphase analysis) is the most common cytogenetic method for biological estimation of radiation absorbed dose. In this method, the unstable chromosomal aberrations (dicentric) in the cultured lymphocytes are scored and converted to absorbed dose. Despite the improvement in the methods and using similar statistical programs in different biodosimetry laboratories, they make different biodosimetry results. The International Atomic Energy Agency has recommended that each laboratory prepare its own dose-response curve. To prepare a curve for the Iran Nuclear Regulatory Authority biodosimetry lab, after obtaining blood samples from 5 volunteers (three men and two women in the age range of 35 to 45 years), the samples were irradiated with different doses (0.1, 0.25, 0.5, 0.75, 1, 2, 3 and 4 Gy) of gamma ray (Cobalt 60). After isolation and culturing of lymphocytes, cultures were treated with colcemid at 45 hours and harvested at 48 hours. After coding and painting of the slides, the dicentric aberrations in the metaphase spreads were scored and then using the Dose Estimate software, the standard dose response curve of human blood lymphocytes for different doses of gamma radiation was depicted. In order to check the curve efficiency, two other samples with unknown radiation doses are scored and the biological dosimetry results compared with the physical doses and the accuracy of dose response curve was confirmed.