MS-H: A novel proteomic approach to isolate and type the E. coli H antigen using membrane filtration and liquid chromatography-tandem mass spectrometry (LC-MS/MS)

Keding Cheng, Mike Drebot, Joanne McCrea, Lorea Peterson, David Lee, Stuart McCorrister, Richard Nickel, Alyssia Gerbasi, Angela Sloan, Debra Janella, Gary Van Domselaar, Daniel Beniac, Tim Booth, Linda Chui, Helen Tabor, Garrett Westmacott, Matthew Gilmour, Gehua Wang

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12 Citations (Scopus)


Serotyping is the long-standing gold standard method to determine E. coli H antigens; however, this method requires a panel of H-antigen specific antibodies and often culture-based induction of the H-antigen flagellar motility. In this study, a rapid and accurate method to isolate and identify the Escherichia coli (E. coli) H flagellar antigen was developed using membrane filtration and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Flagella were isolated from pure culture, digested with trypsin, and then subjected to LC-MS/MS using one of two systems (Agilent-nano-LC-QSTAR XL or Proxeon-nano-LC-LTQ-Orbitrap XL). The resulting peptide sequence data were searched against a custom E. coli flagella/H antigen database. This approach was evaluated using flagella isolated from reference E. coli strains representing all 53 known H antigen types and 41 clinical E. coli strains. The resulting LC-MS/MS classifications of H antigen types (MS-H) were concordant with the known H serogroup for all 53 reference types, and of 41 clinical isolates tested, 38 (92.7%) were concordant with the known H serogroup. MS-H clearly also identified two clinical isolates (4.9%) that were untypeable by serotyping. Notably, successful detection and classification of flagellar antigens with MS-H did not generally require induction of motility, establishing this proteomic approach as more rapid and cost-effective than traditional methods, while providing equitable specificity for typing E. coli H antigens.

Original languageEnglish
Article numbere57339
JournalPLoS One
Issue number2
Publication statusPublished - 21 Feb 2013

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