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Escherichia coli are common pathogens, whereas E. coli O157:H7 is the most notorious E. coli strain, owing to its high virulence that can cause serious adverse effects and death. E. coli contains abundant peroxidases. Thus, the presence of E. coli can be determined by mixing E. coli with its substrate such as 3,5,3′,5′ tetramethylbenzidines (TMB) for endogenous peroxidase reactions. Under the presence of a high concentration of E. coli, colorless TMB turned to bluish, owing to the generation of the complexity of TMB and its oxidized TMB. To further reduce the detectable cell concentration, we developed an affinity-based method combined with an endogenous peroxidase reaction and mass spectrometric detection to detect E. coli. Affinity probes (diameter: ~20 µm) modified with maltose were generated for the enrichment of E. coli from sample solutions. E. coli trapped by the affinity probes was reacted with TMB in the presence of hydrogen peroxide for endogenous peroxidase reactions. Contactless atmospheric pressure ionization mass spectrometry was used for the detection of the reaction product, oxidized TMB (TMB cationic radical), to indicate the presence of target bacteria. The results showed that the developed method can be used to rapidly determine the presence of E. coli from a sample solution based on the detection of the TMB cationic radicals. The lowest detectable concentration of our method against E. coli O157:H7 in buffers and in complex juice samples was as low as ~100 cfu mL−1.

Detection of Escherichia coli by Combining an Affinity-Based Method with Contactless Atmospheric Pressure Ionization Mass Spectrometry