Programmable\nFramework Nucleic Acid-Modified Nanomagnetic\nBeads for Efficient Isolation of Exosomes and Exosomal Proteomics\nAnalysis

Exosomes are increasingly being regarded as emerging\nand promising\nbiomarkers for cancer screening, diagnosis, and therapy. The downstream\nmolecular analyses of exosomes were greatly affected by the isolation\nefficiency from biosamples. Among the current exosome isolation strategies,\naffinity nanomaterials performed comparably better with selectivity\nand specificity. However, these techniques did not take the structure\nand size of exosomes into account, which may lead to a loss of isolation\nefficiency. In this article, a framework nucleic acid was employed\nto prepare a well-designed nanosized bead Fe<sub>3</sub>O<sub>4</sub>@pGMA@DNA TET@Ti<sup>4+</sup> for enrichment of exosomes. The abundant\nphosphate groups in the framework nucleic acid provide binding sites\nto immobilize Ti<sup>4+</sup>, and its rigid three-dimensional skeleton\nmakes them act as roadblocks to barricade exosomes and provide affinity\ninteractions on a three-dimensional scale, resulting in the improvement\nof isolation efficiency. The model exosomes can be effectively isolated\nwith 92% recovery in 5 min. From 100 μL of HeLa cell culture\nsupernatant, 34 proteins out of the top 100 commonly identified exosomal\nproteins were identified from the isolated exosomes by the novel beads,\nwhich is obviously more than that by TiO<sub>2</sub> (19 proteins),\nindicating higher isolation efficiency and exosome purity by Fe<sub>3</sub>O<sub>4</sub>@pGMA@DNA TET@Ti<sup>4+</sup> beads. The nanobeads\nwere finally applied for comparing exosomal proteomics analysis from\nreal clinical serum samples. Twenty-five upregulated and 10 downregulated\nproteins were identified in the lung cancer patients group compared\nto the health donors group, indicating that the novel nanobeads have\ngreat potential in isolation of exosomes for exosomal proteomics analysis\nin cancer screening and diagnosis.