Fig. S1.

Potential sources of SERS hotspots. (A) The center of each 25-μm² area is interrogated by a 1-μm² focused laser beam. (B) Three types of hot spots are possible in the assay: (Left) false positives originating from defects in the film, which are dominated by the MB Raman label, with a strong peak at 1,630 cm⁻¹; (Center) false positives from AuNP aggregates, which are dominated by the NBT Raman label with a strong peak at 1,337 cm⁻¹; and (Right) true-positive protein-mediated binding, indicated by near-equal enhancement of MB and NBT peaks.