Detection of Secretion of Exosomes from Individual Cell in Real-Time by Multifunctional Nanoelectrode-Nanopore Nanopipettes

Abstract

In recent years, exosomes as the intercellular vehicles have received extensive attention for their fundamental roles in biosystem and as novel biomarkers for early detection and therapeutic applications. It has been demonstrated that anti-CD63 can effectively target exosomes from cancer cells. In this work, multifunctional nanoelectrode-nanopore nanopipettes (MNNPs) were prepared to detect exosomes secreted from human cervical cancer cells (HeLa cells) at the single-entity level using the nanopore- based resistive pulse sensing technique. Synthetic liposomes were also measured for comparison. By modifying the anti-CD63 on the carbon nanoelectrode surface near the nanopore, the detected current signals were very different from the signals from the non-modified MNNPs. The differences were induced by the specific interactions between anti-CD63 and the membrane protein CD63 at the surface of exosomes. No obvious differences were observed when liposomes were tested. Furthermore, by utilizing micromanipulation technology, the position of the nanopipette tip could be accurately located, which enabled monitoring the secretion of exosomes from individual cells in a real-time manner. In the control experiment, the probe-to-cell distance was optimized and the HeLa cells incubated with exosomes inhibitor (GW4869) were tested by MNNPs. These significantly reduced translocation signals proved that anti-CD63 modified MNNPs were competent for exosomes detection in complex cellular environments. A glass nanopipette based extracellular exosome detection method was developed. Multifunctional nanopore-nanoelectrodes nanoprobes (MNNPs) were prepared with the modification of antibody at the nanoelectrode surface for the specific detection of exosomes. The focus of this research is that MNNPs can ensure cell viability and detect exosomes secreted from individual human cervical cancer (HeLa) cells in real time by nanopore-based resistance pulse sensing method. The reported results demonstrate that the MNNPs can selectively enrich and detect exosomes in a complex cellular environment, and have the potential to be applied to the study of various nanosale biological entities.