Protein-Enabled\nSize-Selective Defect-Sealing of Atomically\nThin 2D Membranes for Dialysis and Nanoscale Separations

Atomically thin 2D materials present the potential for\nadvancing\nmembrane separations via a combination of high selectivity (from molecular\nsieving) and high permeance (due to atomic thinness). However, the\ncreation of a high density of precise nanopores (narrow-size-distribution)\nover large areas in 2D materials remains challenging, and nonselective\nleakage from nanopore heterogeneity adversely impacts performance.\nHere, we demonstrate protein-enabled size-selective defect sealing\n(PDS) for atomically thin graphene membranes over centimeter scale\nareas by leveraging the size and reactivity of permeating proteins\nto preferentially seal larger nanopores (≥4 nm) while preserving\na significant amount of smaller nanopores (via steric hindrance).\nOur defect-sealed nanoporous atomically thin membranes (NATMs) show\nstability up to ∼35 days during size-selective diffusive separations\nwith a model dialysis biomolecule fluorescein isothiocyanate (FITC)-Ficoll\n70 in phosphate buffer saline (PBS) solution as well as outperform\nstate-of-the-art commercially available dialysis membranes (molecular-weight-cutoff\n∼3.5–5 kDa and ∼8–10 kDa) with significantly\nhigher permeance for smaller solutes KCl (∼0.66 nm) ∼5.1–6\n× 10<sup>–5</sup> ms<sup>–1</sup> and vitamin B12\n(B12, ∼1.5 nm) ∼2.8–4 × 10<sup>–6</sup> ms<sup>–1</sup> compared to small protein lysozyme (Lz, ∼4\nnm) ∼4–6.4 × 10<sup>–8</sup> m s<sup>–1</sup>, thereby allowing unprecedented selectivity for B12/Lz ∼70\nand KCl/Lz ∼1280. Our work introduces proteins as nanoscale\ntools for size-selective defect sealing in atomically thin membranes\nto overcome persistent issues and advance separations for dialysis,\nprotein desalting, small molecule separations/purification, and other\nbioprocesses.