Sandwich fluorimetric method for specific detection of Staphylococcus aureus based on antibiotic-affinity strategy.

A novel antibiotic-affinity strategy was designed for fluorimetric detection of pathogenic bacteria based on the strong affinity of antibiotic agent to the cell wall of bacteria. In this proof-of-concept work, vancocin, a glycopeptide antibiotic for Gram-positive bacteria, was used as a molecular recognition agent to anchor Staphylococcus aureus (S. aureus) cell. To improve the specificity of this method for S. aureus detection, IgG was adopted as the second recognition agent utilizing the binding between Fc region of IgG and S. aureus protein A in the cell wall, to form a sandwich complex. By using fluorescein isothiocyanate as the signal probe, S. aureus whole cells could be directly assayed within a linear range of 1.0 × 10(3)-1.0 × 10(9) CFU mL(-1) with a detection limit of 2.9 × 10(2) CFU mL(-1). The whole assay process could be completed within 130 min when a ready-for-use microplate was adopted. This proposed strategy for pathogenic bacteria detection possessed some attractive characteristics such as high sensitivity, wide linear range, simple manipulation, short assay time, and low cost. Furthermore, this sandwich mode also showed ideal specificity because vancocin and IgG bound with S. aureus at two distinct sites. It opened up a new pathway for high-throughput screening of pathogenic bacteria in medical diagnosis, food safety, bioterrorism defense, and drug discovery.