Breast cancer is the most common cancer and the second cause of cancer-related death in women, especially in the age of 20-59 years. It is very important to diagnose it in the early stages of development due to high chance of survival. In this research, the early detection of two microRNAs involved in breast cancer including miR-21 and miR-155 was performed simultaneously using a nanobiosensor based on a special G-quadruplex structure and a colorimetric manner. This nanobiosensor contains two probes (p1, p2) that play a role in the formation of a special structure called DNA-G4. This structure has peroxidase-like properties and can oxidize TMB and produce a blue color. The diagnostic method is designed as a signal off, where the hybridization of probes with microRNA sequences, no DNA-G4 structure is formed and no color change is observed. The results of this study showed that the linear range of response is in the range of 2 to 10 nm and limit of detection in buffer, blood and urine samples was calculated as 0.43 nM, 0.54 nM, and 0.62 nM (R = 0.98), respectively. Evaluation using the method for cancer monitoring can be a simple, fast and cost-effective technique.