Neuroblastoma (NB) is an aggressive pediatric cancer originating from neural crest cells and is often associated with poor prognosis due to genetic alterations such as MYCN amplification and 11q deletion. MicroRNAs (miRNAs) are short RNA molecules that influence gene regulation and play significant roles in tumor formation and progression. This analysis considers the expression of hsa-miR-708-5p and hsa-miR-708-3p in five neuroblastoma cell lines (SH-SY-5Y,NB69, IMR32, SK-N-AS, and SK-N-BE), with and without the above mentioned genetic alterations. Quantitative PCR (qPCR) was used to measure miRNA expression, with U6B small nuclear RNA (RNU6B) serving as an endogenous control. In this study, miR-708-5p was detected in all neuroblastoma cell lines, while miR-708-3p expression was observed only in NB69, SH-SY-5Y, and IMR32. The 11q deletion in SK-N-AS may be the possible reason for the absence of miR-708-3p, although miR-708-5p was still expressed, likely due to a heterozygous deletion that left intact copies of the gene. In SK-N-BE, the lack of miR-708-3p expression may result from epigenetic silencing, chromosomal alterations affecting the miRNA locus, or natural absence of expression under the studied conditions. Statistical analysis confirmed significant expression differences in both miRNAs. ANOVA showed lower miR-708-5p expression in the 11q-deleted group, while Welch’s t-test revealed reduced miR-708-3p expression in IMR-32 versus control, indicating biologically relevant variation. Expanding future studies with larger sample sizes across different cell lines is necessary to better understand the complex regulation of miR-708-5p and miR-708-3p in neuroblastoma and to explore their potential as biomarkers or therapeutic targets.