Affinity enrichment of placental extracellular vesicles from minimally processed maternal plasma with magnetic nanowires

Affinity based enrichment of cell/tissue specific extracellular vesicles (EVs) with magnetic materials and analysis of their molecular cargo has the potential to improve assay sensitivity/specificity compared to whole plasma analysis. For example, syncytiotrophoblast EVs (STBEVs) shed from the placenta during pregnancy carry placental diagnostic markers relevant to pregnancy complications linked to placental insufficiency such as placental alkaline phosphatase (PLAP), Neprilysin (NEP) and Placental Protein 13 (PP13). However, the need for sample pre-enrichment of EVs from plasma adds significant complexity, time and cost. We report an affinity-based cell/tissue specific EV enrichment direct from plasma based on iron-oxide magnetic nanowires (NWs) coated with reversible-addition-fragmentation-chain-transfer (RAFT) polymers and conjugated with anti-PLAP antibodies. As anticipated the complex protein environment of minimally processed plasma significantly decreased STBEV enrichment yield. However, an optimized RAFT polymeric coating successfully mitigated the detrimental effect of the protein corona, yielding significantly improved STBEV recovery compared to Dynabeads™ in unenriched diluted plasma. Despite the presence of significant soluble PLAP protein, STBEV enrichment could be performed directly from the plasma of pregnant women (including preeclamptic samples) within 1.5 hours, enabling quantification of two placental protein markers PP13 and NEP with known diagnostic relevance to preeclampsia. Direct affinity-enrichment of STBEVs with high performance magnetic materials has the potential to underpin rapid clinical diagnostic assays for preeclampsia and related pregnancy complications.