This study aimed to to evaluate the stability of commonly used endogenous control genes for messenger RNA (mRNA) (N = 16) and miRNAs (N = 3) expression studies in prostate cell lines following irradiation. The stability of endogenous control genes expression in irradiated (6 Gy) versus unirradiated controls was quantified using NormFinder and coefficient of variation analyses. HPRT1 and 18S were identified as most and least stable endogenous controls, respectively, for mRNA expression studies in irradiated prostate cells. SNORD48 and miR16 miRNA endogenous controls tested were associated with low coefficient of variations following irradiation (6 Gy). This study highlights that commonly used endogenous controls can be responsive to radiation and validation is required prior to gene/miRNAs expression studies.