Clearing for insight: A light-sheet microscopy workflow for 3D vascular reconstruction in human first trimester placental villi and decidua.

The 3D visualization of the maternal-fetal interface is essential for understanding human placental development, yet remains technically challenging due to tissue opacity and structural complexity. Here, we describe a robust light-sheet fluorescence microscopy workflow optimized for imaging first-trimester placental villi and decidua, which were cleared, multiplex immunofluorescence labeled, and imaged by light-sheet microscopy. Pan-cytokeratin, CD31 or CD34, and alpha smooth muscle actin staining enabled 3D reconstruction of the vascular architecture in placental villi and decidua parietalis. This workflow may aid in elevating previous 2D observations into a 3D context, offering a more comprehensive view of tissue organization at the early maternal-fetal interface.