Prokaryotic expression of Stx1B subunit of Escherichia coli O157:H7 used to generate monoclonal antibody.
The Shiga-like toxins (Stx) are critical virulence factors for enterohemorrhagic Escherichia coli (EHEC), causing severe human illnesses of bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome (HUS). A subunit of Stx, Stx1B protein, mediates the interaction between EHEC and the specific receptor of host organs. In this article, the recombinant expression vector pGEX-Stx1B bearing a signal peptide sequence-deleted Stx1B gene was constructed and the expression of protein of interest was achieved in a prokaryotic system. The resulting Stx1B protein was used to immunize BALB/c mice followed by the preparation of corresponding monoclonal antibodies (MAbs). One MAb (1G11) was generated. Western blot analysis showed that the MAb was capable of reacting with EHEC Stx1B protein. It had high affinity binding to the Stx1B protein and it distinguished EHEC from other control bacteria. Therefore, the MAb generated in this study can be used as a specific reagent to investigate the pathogenesis mechanism and to develop a diagnostic method of EHEC infection in both humans and animals.
Li
Guangxing
G
College of Veterinary Medicine, Northeast Agricultural University, Harbin, China.
Hong
Jiehua
J
Ren
Xiaofeng
X
Yin
Jiechao
J
Feng
Shuzhang
S
Huo
Guicheng
G
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D019453:Escherichia coli O157 / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D006801:Humans; D006825:Hybridomas; D007114:Immunization; D051379:Mice; D008807:Mice, Inbred BALB C; D021122:Protein Subunits; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D022622:Shiga Toxin 1 / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D021122:Protein Subunits; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D022622:Shiga Toxin 1