Background MicroRNAs (miRs), which are stable in the blood, comprise small non‐coding RNAs that regulate gene expression. They have important roles in almost all biological pathways, especially in cancer‐relevant processes, and have an abnormal expression in breast cancer. In recent studies, the aberrant expression level of various microRNAs has been demonstrated in human cancer. In the present study, the status of serum microRNA‐210‐3p and microRNA‐660‐5p expression levels in breast cancer patients was determined compared to healthy controls. Methods Serum samples were collected from 40 newly diagnosed breast cancer patients and 40 healthy controls. A real‐time quantitative polymerase chain reaction was utilized to detect the expression levels of these microRNAs. Data analysis was conducted with p < 0.05 being considered statistically significant. Results The data obtained showed that serum levels of miR‐660‐5p and miR‐210‐3p were significantly up‐regulated in breast cancer patients compared to healthy controls (p < 0.001 and p = 0.001, respectively). In addition, significant up‐regulation was observed in the early stage (in situ, stage I and II) of breast cancer patients (n = 25) compared to healthy (n = 40) controls (p < 0.001 and p < 0.05, respectively). Receiver‐operating characteristic curve analysis indicated that the serum miR‐660‐5p and miR‐210‐3p levels have reasonable sensitivity (79% and 68%) and specificity (61% and 51%) for the detection of breast cancer patients (area under the receiver‐operating curve = 0.774 and 0.716, respectively). Conclusions Although the results show a reasonable diagnostic accuracy of these microRNAs for detection of breast cancer in this small and preliminary study, further large‐scale studies are essential to confirm the presented results.