New Stx2e Monoclonal Antibodies for Immunological Detection and Distinction of Stx2 Subtypes

PLoS One. 2015 Jul 20;10(7):e0132419. doi: 10.1371/journal.pone.0132419. eCollection 2015.

Abstract

Background: Stx2e is a primary virulence factor in STEC strains that cause edema disease in neonatal piglets. Though Stx2a and Stx2e are similar, many antibody-based Stx detection kits are designed to detect Stx2a and do not recognize the Stx2e subtype.

Methods and findings: Four monoclonal antibodies against Stx2e were developed and characterized. Two of these mAbs recognize the B subunit of Stx2e, Stx2f, and to a lesser extent, Stx2b, Stx2c, and Stx2d. The other two mAbs recognize the A subunit of Stx2e, and cross-react with all Stx2 subtypes except Stx2f. The most sensitive sandwich ELISA using these mAbs has a limit of detection for Stx2e of 11.8 pg/mL. The ability of the neutralizing antibody Stx2e-2 to block Stx2e-receptor binding in Vero cells was visualized using immunofluorescence. Combinations of these and previously developed mAbs permit ELISA-based differentiation between closely related Stx2a, Stx2c, and Stx2d (using mAbs Stx2-5/2-1, Stx2-5/2e-2, and Stx2e-3/2e-2, respectively).

Conclusions: The sensitive immunoassays developed in this study should augment our capacity to detect Stx2e in porcine environments and biological samples. Moreover, immunoassays that can distinguish between the closely related Stx2a, Stx2c, and Stx2d subtypes can be useful in quickly analyzing Stx subtypes in samples containing more than one strain of STEC.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Antibodies, Monoclonal*
  • Chlorocebus aethiops
  • Enzyme-Linked Immunosorbent Assay
  • Escherichia coli Infections / diagnosis*
  • Escherichia coli Infections / metabolism
  • Mice
  • Shiga Toxin 2 / metabolism*
  • Vero Cells

Substances

  • Antibodies, Monoclonal
  • Shiga Toxin 2

Grants and funding

This study was funded by the United States Department of Agriculture-Agricultural Research Service (USDA-ARS) National Program NP108 Current Research Information System (CRIS) project 5325-42000-048-00D. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.