All-in-One Strategy for Downstream Molecular Profiling of Tumor-Derived Exosomes.

In light of the significance of exosomes in cancer diagnosis and treatment, it is important to understand the components and functions of exosomes. Herein, an all-in-one strategy has been proposed for comprehensive characterization of exosomal proteins based on nanoporous TiO clusters acting as both an extractor for exosome isolation and a nanoreactor for downstream molecular profiling. With the improved hydrophilicity and inherent properties of TiO, exosomes can be captured by a versatile nanodevice through the specific binding and hydrophilicity interaction synergistically. The strong concerted effect between exosomes and nanodevices ensured high efficiency and specificity of exosome isolation with high recovery and low contaminations. Meanwhile, highly efficient downstream proteomic analysis of the purified exosomes was also enabled by the nanoporous TiO clusters. Benefiting from the porous structure of the nanodevice, the lysed exosomal proteins are highly concentrated in the nanopore to achieve high-efficiency in situ proteolytic digestion. Therefore, the unique features of the TiO clusters ensured that all the complex steps about isolation and analysis of exosomes were completed efficiently in one simple nanodevice. The concept was first proved with exosomes from cell culture medium, where a high number of identified total proteins and protein groups in exosomes were obtained. Taking advantage of these attractive merits, the first example of the integrated platform has been successfully applied to the analysis of exosomes in complex real-case samples. Not only 196 differential protein biomarker candidates were discovered, but also many more significant cellular components and functions related to gastric cancer were found. These results suggest that the nanoporous TiO cluster-based all-in-one strategy can serve as a simple, cost-effective, and integrated platform to facilitate comprehensive analysis of exosomes. Such an approach will provide a valuable tool for the study of exosome markers and their functions.