Programmable Framework Nucleic Acid-Modified Nanomagnetic Beads for Efficient Isolation of Exosomes and Exosomal Proteomics Analysis.

ABSTRACT

Exosomes are increasingly being regarded as emerging and promising biomarkers for cancer screening, diagnosis, and therapy. The downstream molecular analyses of exosomes were greatly affected by the isolation efficiency from biosamples. Among the current exosome isolation strategies, affinity nanomaterials performed comparably better with selectivity and specificity. However, these techniques did not take the structure and size of exosomes into account, which may lead to a loss of isolation efficiency. In this article, a framework nucleic acid was employed to prepare a well-designed nanosized bead Fe3O4@pGMA@DNA TET@Ti4+ for enrichment of exosomes. The abundant phosphate groups in the framework nucleic acid provide binding sites to immobilize Ti4+, and its rigid three-dimensional skeleton makes them act as roadblocks to barricade exosomes and provide affinity interactions on a three-dimensional scale, resulting in the improvement of isolation efficiency. The model exosomes can be effectively isolated with 92% recovery in 5 min. From 100 µL of HeLa cell culture supernatant, 34 proteins out of the top 100 commonly identified exosomal proteins were identified from the isolated exosomes by the novel beads, which is obviously more than that by TiO2 (19 proteins), indicating higher isolation efficiency and exosome purity by Fe3O4@pGMA@DNA TET@Ti4+ beads. The nanobeads were finally applied for comparing exosomal proteomics analysis from real clinical serum samples. Twenty-five upregulated and 10 downregulated proteins were identified in the lung cancer patients group compared to the health donors group, indicating that the novel nanobeads have great potential in isolation of exosomes for exosomal proteomics analysis in cancer screening and diagnosis.