Hitting Two Birds with One Stone: Magnetism–Fluorescence Janus Composites for Simultaneous Targeting of Serum Exosomes and Trapping of Phosphoproteins

Designing a highly efficient serum exosome (sEV) phosphorylation proteomics platform with specific sEV targeting and phosphoprotein trapping for biomarker identification in renal cell carcinoma (RCC) is challenging. Herein, we fabricated a Janus composite serving as one stone for two birds, enabling directed localization of sEVs and phosphoprotein enrichment, thus realizing the trapping of differential sEV phosphoproteins. Rhodamine B (RhB) and guanidine (Gua) endowed the composite, denoted as Janus@RhB@Gua, with magnetism–fluorescence dual functionality. The material possessed high sEV uptake efficiency (14000 μg/mL) and exhibited high sensitivity (with a detection limit of 0.1 fmol/μL) for phosphopeptide detection coupled with liquid chromatography–tandem mass spectrometry (LC-MS/MS). Furthermore, we confirmed its effectiveness in the control experiment, wherein 4095 phosphopeptides (1802 N-phosphopeptides and 2293 O-phosphopeptides) corresponding to 986 phosphoproteins were enriched from RCC patient (n = 6) and normal control (n = 3) serum samples. Twelve differentially sEV phosphoproteins were identified and confirmed as playing a crucial role in the pathogenic pathway of RCC. Overall, these findings highlight the dual functionality of Janus@RhB@Gua, which enables effective sEV phosphoprotein trapping, thereby facilitating the identification of RCC biomarkers.