Gold Electron Microscopy Grids with Anisotropic Foil Geometry Enable On-Grid Contact Guidance.

All-gold electron microscopy (EM) grids reduce beam-induced motion relative to conventional holey carbon supports and provide biocompatible substrates for cellular cryo-EM. However, placing customizable all-gold grid fabrication in the hands of researchers requires accessible processes based on standard microfabrication tools. We report a wafer-scale process using microfabrication techniques available in most academic cleanrooms, such as lift-off metallization, electroplating, and sacrificial layer release, to fabricate 594 all-gold grids per 4-inch wafer without individual grid handling. A numerical electroplating model provides a quantitative framework to relate gold deposition, grid-bar thickness, and tilt-compatible grid geometry. We show that oval 2 μm × 6 μm foil holes bias on-grid actin organization by substrate geometry alone, without chemical micropatterning. The EM grids supported a 2.15 Å apoferritin single-particle reconstruction on a 200 kV cryo-TEM and are compatible with protein micropatterning and cryo-electron tomography of cells. This platform establishes an accessible route to programmable, application-specific all-gold cryo-EM supports that couple high-resolution structural imaging with engineered control of cellular organization.