Xiang Gu (Shiitake) is a culinary mushroom and Chinese herbal medicine. Xiang Gu strengthens the Spleen, benefits Qi, eliminates pathogens and regulates Yin and Yang. In numerous studies, lentinan, a polysaccharide isolated from Xiang Gu, has been shown to have immunomodulary effects. The hypothesis of the study was that lentinan, at optimum doses, would proliferate chicken lymphocytes in-vitro cell cultures. Eleven concentrations of lentinan were added to 4 wells of lymphocyte cell cultures each. Following incubation and processing, the absorbance was measured at 570nm (A570 values) with an ELISA plate reader to determine non-toxic and optimum concentrations of lentinan, compared to a cell control group. From these data, 5 concentrations of lentinan were selected to study the effect on lymphocytes, alone or combined with the T-lymphocyte mitogen phytohemagglutinin (PHA). The mean ± standard deviation (M±SD) A570 values of Groups 3-5 with 37.5, 18.75 and 9.375 μg/mL respectively of lentinan alone were significantly higher (p<0.05) than all other groups of lentinan alone and the cell control group. The M±SD A570 values of Groups 3-5 with 37.5, 18.75 and 9.375 μg/mL respectively of lentinan combined with PHA were significantly higher (p<0.05) than all other groups receiving lentinan and PHA and a PHA control group. From the results of this study, the hypothesis was accepted since 37.5, 18.75 and 9.375 μg/mL lentinan significantly produced lymphocyte proliferation alone and synergistically with PHA. This study provides a theoretical basis for further in-vivo studies of lentinan as an immune-enhancing treatment for birds and animals.