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Simple Summary In the last several years, the animal production sector has been very concerned about the overuse of antibiotics and the concomitant increase in antimicrobial resistance. The objective of this paper was the evaluation of the immune properties of different extracts isolated from Chlorella sorokiniana applied to sheep peripheral blood mononuclear cells (PBMCs). In particular, the effects of the unsaponified fraction (UP), the acetylated unsaponified fraction (AUP), and the total lipid fraction (TL) at two concentrations (0.4 and 0.8 mg/mL) on the proliferative response of sheep cells and their cytokine secretion were tested. All the extracts showed an inhibitory effect on cell proliferation, particularly in the presence of the UP fraction at a 0.4 mg/mL concentration. As regards cytokine production, the UP extract tested at a 0.8 mg/mL concentration increased interleukin(IL)-10 production, whereas the TL fraction at 0.4 mg/mL showed a cytokine profile characterized by an increase in both IL-10 secretion and the anti-inflammatory cytokine IL-6, and, to a lesser extent, IL-1β secretions, which are proinflammatory cytokines. These results supported the hypothesis that extracts from Chlorella sorokiniana could be considered useful ingredients to be integrated into animal feed with the aim to control immune responses during inflammation and minimize the use of antibiotics. Abstract The objective of this experiment was to study the effects of the unsaponified fraction (UP), the acetylated unsaponified fraction (AUP), and the total lipid fraction (TL) extracted and purified from Chlorella sorokiniana (CS) on the proliferation and cytokine profile of sheep peripheral blood mononuclear cells (PBMCs). Cells were cultured with 0.4 mg/mL and 0.8 mg/mL concentrations of each extract (UP, AUP, and TL fractions) and activated with 5 μg/mL concanavalin A (ConA) and 1 μg/mL lipopolysaccharide (LPS) at 37 °C for 24 h. PBMCs cultured with ConA and LPS represented the stimulated cells (SC), and PBMCs without ConA and LPS represented the unstimulated cells (USC). Cell-free supernatants were collected to determine IL-10, IL-1β, and IL-6 secretions; on cells, measurement of proliferation was performed. All the extracts tested significantly decreased the cell proliferation; in particular, the UP fraction at 0.4 mg/mL showed the lowest proliferative response. Furthermore, at 0.8 mg/mL, the UP fraction enhanced IL-10 secretion. On the contrary, the TL fraction at 0.4 mg/mL induced an increase in IL-10, IL-6, and, to a lesser extent, IL-1β secretions by cells. The AUP fraction did not change cytokine secretion. The results demonstrated that CS extracts could be useful ingredients in animal feed in order to minimize the use of antibiotics by modulating cell proliferation and cytokine response.