Acute lymphoblastic leukemia (ALL) is a malignant disease caused by the abnormal growth and development of early non-granular white blood cells, or lymphoblasts. ALL occurs predominantly in children, peaking at 4 years of age. Interferons (IFNs) are the biological agents involved in the antiviral responses and the inhibition of tumor growth. In oncology, the IFNs are an important treatment options for a number of solid tumors and hematological malignancies. The IFN-β1gene consists of 840 bp and it lacks intron sequences. Seventy patients suffering from Childhood Acute Lymphoblastic Leukemia were studied and analyzed for mutation analysis of IFN-β1. So, 58.3% of the patients suffering from ALL were observed and compared with 41.7% normal DNA samples as control. PCR was performed and amplified gene products were detected on agarose gel and PAGE. Finally, SSCP was performed and mobility shift was observed in 7 out of 70 ALL samples. We observed that ALL disease typically develops in children ages 1-10 years old and is more prevalent in males than females. Mutation in the IFN-β1 gene was significantly related to 1-5 years in case of association of mutation with age.