To assess the relationship between MYCN amplification in neuroblastoma (NB), tumor stage and prognosis; and to evaluate the usefulness of CD44 in predicting prognosis of NB.Differential polymerase chain reaction (D-PCR) with serial dilution assay was used to quantify the MYCN gene copy number in 33 paraffin-embedded tissue samples of NB. All the samples were also studied by immunohistochemistry for CD44. The results were correlated with various prognostic factors of NB, including patient age, tumor stage, pathologic type and MYCN gene amplification.MYCN amplification was identified in 10 of the 33 samples studied (30.3%), which all were in high clinical stage (stage III or IV) and occurred in patients older than 1 year of age. MYCN amplification also significantly correlated with poor clinical outcome (P < 0.01). CD44 was positive in 21 cases and often occurred in patients below 1 year of age, in low clinical stage, with favorable histology and without MYCN gene amplification. The two-year survival rate of CD44-positive group (57.1%) was higher than that of CD44-negative group (8.3%, P < 0.01). Stronger CD44 expression was also associated with better prognosis (P < 0.01).MYCN gene amplification is significantly associated with advanced tumor stage and poor prognosis in patients with NB. CD44 expression is a reliable marker for better prognosis and is complementary to MYCN amplification assay. D-PCR with serial dilution assay is also suitable for clinical use in quantifying MYCN copy number in NB.