Biological metamaterials with a specific size and spacing are necessary for developing highly sensitive and selective sensing systems to detect hazardous bacteria in complex solutions. Herein, the construction of peptidoglycan-binding protein (PGBP)-based metamaterials to selectively capture Gram-positive cells with high efficacy is reported. Nanoimprint lithography was used to generate a nanohole pattern as a template, the inside of which was modified with nickel(II)-nitrilotriacetic acid (Ni-NTA). Then, PGBP metamaterials were fabricated by immobilizing PGBP via chelation between Ni-NTA and six histidines on PGBP. Compared to the flat and spread PGBP-covered bare substrates, the PGBP-based metamaterials enabled selective capturing of Gram-positive bacteria with high efficacy, owing to enhanced interactions between the metamaterials and bacterial surface not shown in bulk materials. Thereafter, the specific strain and quantitative information of the captured bacteria was obtained by surface-enhanced Raman scattering mapping analysis in the 1 to 1 × 106 cfu/mL range within 30 min. It should be noted that no additional signal amplification process was required for lowly abundant bacteria, even at the single-bacterium level. The PGBP-based metamaterials could be regenerated multiple times with preserved sensing efficiency. Finally, this assay can detect specific Gram-positive bacteria, such as Staphylococcus aureus, in human plasma.
Keywords: bacteria; bacteria capturing; nanoImprint lithography; peptidoglycan-binding protein; protein-based metamaterials; sensing; surface-enhanced Raman scattering.