Human leukocyte antigen G (HLA-G) is a non-classical major histocompatibility class Ib antigen with multiple immune regulatory functions including the induction of immune tolerance in malignancies. The goal of our study was to investigate the expression of membrane form of HLA-G in acute lymphoblastic leukemia (ALL) before and after therapy in a trial to evaluate its role as a tumor escape mechanism and prognosis. So we measured its expression by reverse transcription (RT)-PCR in peripheral blood mononuclear cells of 25 (ALL) patients and 15 healthy controls and correlated our findings with a variety of clinical and laboratory variables and two important cytokines, IL-10 and INF-γ, and with natural killer (NK) cells. Serum levels of IL-10 and INF-γ were measured by ELISA. NK cells were quantitated by flow cytometry. The best cutoff values for the investigated markers were determined by ROC curve. The current study showed that membrane-bound HLA-G expression levels and positivity rates above the cutoff value 0.37 were significantly higher in ALL patients at diagnosis compared to after therapy and both showed significant higher levels than in normal control group (P < 0.01). Moreover, IL-10 and INF-γ serum levels were significantly elevated in ALL patients at time of diagnosis compared to healthy controls with a significant reduction in their levels in ALL patients after receiving chemotherapy. Membrane HLA-G expression showed a significant positive correlation with lactate dehydrogenase, peripheral and bone marrow blast cells and with IL-10 and INF-γ. The positive correlation of membrane HLA-G expression with both IL-10 and INF-γ serum levels supports the speculation that both cytokines may be involved in the control of HLA-G expression. HLA-G showed a negative correlation with NK cells confirming its importance in tumor escape through down-regulation of NK cells. In conclusion, HLA-G expression could be used as a prognostic tumor marker to monitor disease state and improvement in ALL.