Current protocols for production of monocyte-derived dendritic cell (moDC) suitable for immunotherapy are obtained from monocytes incubated with IL-4 and GM-CSF in addition with different combinations of pro-inflammatory cytokines. Recently it has been described a new combination of cytokines (including type I and type II interferons) to induce Type I polarized alpha DC (aDCs), characterized by high production of IL-12p70. MoDCs process usually takes 7 days of incubation with either of these cytokine combinations. We compare the maturation phenotype and in vitro functionality of two-day derived DCs using the standard (2d-stDCs) and alpha DCs (2d-aDCs) cytokine combinations. 2d-aDCs or 2d-stDCs were maturated with cytokine cocktails for 24h. DCs phenotype was measured by flow cytometry (FC) with anti-CD80, CD83, CCR7, CD123, CD11c, HLA-DR, CD209 and CD14, and cytokine secretion was measured in DCs supernatant by CBA. For in vitro activation of CD4 (LTBI donor) or CD8 T cells (HER-2/neu positive breast cancer patient) were co-cultured with fresh or frozen DCs pulsed with peptides derived from Mycobacterium tuberculosis (M.tb.) for CD4 activation or HLA-A*02:01 restricted HER-2/neu or NY-ESO1 peptides for CD8 activation. Memory sub-populations, activation markers and tetramer staining was determined by FC. 2d-stDCs and 2d-aDCs have very similar maturation markers with an increase of CD80, CD83, HLA-DR, and a decrease of CD209 and CD14. As expected IL-12p70 is only secreted by 2d-aDCs. 2d-aDCs or 2d-stDCs can induce IFN-¿+ CD4+ T cells and secretion of TNF-¿ in response to M. tb antigens, despite of the use of fresh or frozen DCs. Finally chemotherapy affects peptide specific memory sub-populations increasing IFN-¿ and TNF-¿ secretion in CD8 T cells. Altogether, these results suggest that 2d-aDCs are functional are suitable to monitoring the immunogenicity of tumors in response to chemotherapy in breast cancer patients.