Circulating cancer markers are metabolic products found in body fluids of cancer patients, which are specific for a certain type of malignant tumors. Cancer marker detection plays a key role in cancer diagnosis, treatment, and disease monitoring. The growing need for early cancer diagnosis requires quick and sensitive analytical approaches to detection of cancer markers. The approach based on the photonic crystal surface mode (PC SM) detection has been developed as a label-free high-precision biosensing technique. It allows real-time monitoring of molecular and cellular interactions using independent recording of the total internal reflection angle and the excitation angle of the PC surface wave. We used the PC SM technique for simultaneous detection of the ovarian cancer marker cancer antigen 125 and two breast cancer markers, human epidermal growth factor receptor 2 and cancer antigen 15-3. The new assay is based on the real-time flow detection of specific interaction between the antigens and capture antibodies. Its particular advantage is the possibility of multichannel recording with the same chip, which can be used for multiplexed detection of several cancer markers in a single experiment. The developed approach demonstrates high specificity and sensitivity for detection of all three biomarkers.