This study was designed to prepare and evaluate the specific Mycobacterium bovis (M. bovis) Bacillus Calmette-Guerin (BCG) transfer factor (TF) from spleen cells of two lambs vaccinated subcutaneously S/C with BCG vaccine twice 2 weeks' intervals, similarly one lamb injected with normal saline used as a donor of non-specific TF. The lambs were clinically examined at zero time and along 3 days post each immunization, then, delayed type hypersensitivity (DTH) was checked post 4 weeks. TF was obtained from spleen cells of the lambs, inspected for sterility; safety.TF was evaluated in 4 groups (five in each) of lambs 3-4 months old. The first group (recipient) was injected S/C with 2 ml of specific BCG-TF, similarly the 2nd, 3rd and 4th groups were injected with nonspecific TF, BCG and normal saline respectively. The clinical signs and DTH test were checked as described above, only the specific TF recipient and BCG vaccinated lambs gave a positive DTH test. The serum concentration of inerlukin-17 (IL-17), interferongamma (IFN-γ) and migration inhibitory factor (MIF) cytokines were measured by Enzyme-linked immunosorbent assay (ELISA), which revealed a highly significant increase in the TF recipient lambs in comparison with the control and vaccinated groups. It concluded that the prepared specific M. bovis-BCG transfer factor was effective to induce cell mediated immunity via DTH and increasing the serum concentration of cytokines in recipient lambs.