Ovarian cancer (OC) is a lethal disease diagnosed at advanced stages due to the lack of specific biomarkers. Tyrosine receptor kinase B (TrkB), which has recently been found to be related to OC progression, represents a promising potential biomarker for OC diagnosis and prognosis. The discovery of circulating exosomes as biomarkers for various diseases led us to explore exosomal TrkB in OC. Our previous study proved that the expression of TrkB was elevated in OC tissues. In this study, we focused on the detection of exosomal TrkB in OC. Exosomes were first gathered from three different OC cell lines' conditioned medium, serum samples of patients with OC as well as xenograft mice serum by serial centrifugation method. Then, we identified exosomes by transmission electron microscopy, NanoSight analysis, and expression of typical exosomal protein markers. The existence of TrkB in exosomes was measured by Western blot analysis, and the expression was detected by enzyme-linked immunosorbent assay. In this study, we demonstrated that exosomes could derive from OC cell lines, serum from OC xenograft nude mice, and clinical patients. Our study shows that serum exosomal TrkB may be considered a minimally invasive biomarker for OC.