Nigella sativa has been used since ancient time for curing various infectious diseases due to its miraculous biochemical and pharmacological properties. It belongs to the Ranunculaceae family and generally used as food additives and also known as black cumin. The active components in black cumin have shown remarkable characteristics like antimicrobial, anti-diabetics, anti-cancerous, anti-tumoral, analgesic, anti-fungal, anti-inflammatory, and immunological and immune-potentiating effects. In the present study, we investigated the immunomodulatory effect of Nigella sativa seeds extract on the chicken peripheral blood mononuclear cells (PBMCs). The chicken PBMCs were stimulated with two different doses (125 μg/ml; 250 μg/ml) of Nigella sativa seeds extract and the cells were harvested at different time points till 48 h post-stimulation for analysis of various immune gene expression by quantitative real-time PCR. High dose (250 μg/ml) of Nigella sativa seed extract showed remarkable induction of immune response genes (IL-1β, IFN-β, IFN-γ, IL-4, IL-10, IL-13, IL-12 transcripts) expression at 48h post-stimulation. The results indicated the immunomodulatory activity of the Nigella sativa seed extract on the chicken PBMCs and suggested the potential use as an adjuvant with various vaccines in chicken.