Abstract
Bulk methods to fractionate organelles lack the resolution to capture single-cell heterogeneity. While microfluidic approaches attempt to fractionate organelles at the cellular level, they fail to map each organelle back to its cell of origin—crucial for multiomics applications. To address this, we developed VacTrap, a high-throughput microfluidic device for isolating and spatially indexing single nuclei from mammalian cells. VacTrap consists of three aligned layers: (1) a Bis-gel microwells layer with a ‘trapdoors’ (BAC-gel) base, fabricated atop a through-hole glass slide; (2) a PDMS microwell layer to receive transferred nuclei; and (3) a vacuum manifold. VacTrap operation begins with cell lysis using DDF to release intact nuclei into the Bis-gel microwells, while cytoplasmic proteins are electrophoresed into the Bis-gel. Upon exposure to DTT and vacuum force, the trapdoors open, allowing nuclei to transfer to the PDMS microwells. VacTrap dissolves the trapdoors within 3-5 minutes and achieve synchronized nuclei transfer with 98% efficiency across 80% of trapdoors in a 256-microwell array, surpassing the <1% efficiency of passive transfer without vacuum. Morphology analysis confirmed preservation of organelle integrity throughout VacTrap operation. By enabling spatial indexing of nuclei back to their original cell, VacTrap provides a robust, high-throughput tool for single-cell multiomics applications.
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