Label-Free Detection and Discrimination of Bacterial Pathogens Based on Hemin Recognition

Hemin linked to hexa(ethylene glycol)bishydrazide was patterned by inkjet printing into periodic microarrays, and evaluated for their ability to capture bacterial pathogens expressing various hemin receptors. Bacterial adhesion was imaged under darkfield conditions with Fourier analysis, supporting a label-free method of pathogen detection. Hemin microarrays were screened against a panel of 16 bacteria and found capable of capturing multiple species, some with limits of detection as low as 10(3) cfu/mL. Several Gram-positive strains including Staphylococcus aureus and Bacillus anthracis also exhibited rapid adhesion, enabling pattern recognition within minutes of exposure. This can be attributed to differences in hemin acquisition systems: aggressively adherent bacteria express cell-surface hemin receptors (CSHRs) that enable direct hemin binding and uptake, whereas other types of bacteria including most Gram-negative strains rely on the secretion and recapture of soluble proteins (hemophores) for hemin acquisition, with consequently longer times for ligand binding and detection.