Fabrication of Thiolated Protein A Sensors

BK-7 glass microscope slides (Corning, Painted Post, NY, USA) were cleaned with a boiling piranha solution (3:1 H₂SO₄ and 30% H₂O₂) for 2 h and then subsequently rinsed with water and absolute ethanol before drying under compressed air. A 2 nm layer of chromium (0.5 Å s⁻¹) followed by a 50 nm layer of gold (1.0 Å s⁻¹) was deposited onto the glass slides using electron-beam evaporation (Temescal, Berkeley, CA, USA) at 5 × 10⁻⁶ Torr in a Class 1000 cleanroom facility (UCR Center for Nanoscale Science & Engineering). These gold sensor chips were rinsed with ethanol and dried under compressed air before being coated with 10 μg mL⁻¹ thiolated protein A (Protein Mods, Madison, WI, USA) in 1× PBS for 2 h at room temperature (ca. 23 °C), followed by a 1 mM solution of 3-mercapto-1-propanol (MPO; Millipore-Sigma, St. Louis, MO, USA) in 1× PBS for 1 h at room temperature. The sensor chips were rinsed with nanopure water (≥18 MΩ·cm, Barnstead E-Pure) and dried under a nitrogen stream after each step before final storage at 4 °C. Nanoglassified gold sensor chips were prepared by depositing a thin layer of silica (ca. 2–4 nm) via plasma-enhanced chemical vapor deposition using a Unaxis Plasmatherm 790 (Santa Clara, CA, USA) directly after electron-beam evaporation of gold, and these chips were not subject to any further functionalization.^{27 (link)}