Identification of immunogenic proteins and generation of antibodies against Salmonella Typhimurium using phage display.
BACKGROUND
Solely in Europoe, Salmonella Typhimurium causes more than 100,000 infections per year. Improved detection of livestock colonised with S. Typhimurium is necessary to prevent foodborne diseases. Currently, commercially available ELISA assays are based on a mixture of O-antigens (LPS) or total cell lysate of Salmonella and are hampered by cross-reaction. The identification of novel immunogenic proteins would be useful to develop ELISA based diagnostic assays with a higher specificity.


RESULTS
A phage display library of the entire Salmonella Typhimurium genome was constructed and 47 immunogenic oligopeptides were identified using a pool of convalescent sera from pigs infected with Salmonella Typhimurium. The corresponding complete genes of seven of the identified oligopeptids were cloned. Five of them were produced in E. coli. The immunogenic character of these antigens was validated with sera from pigs infeced with S. Tyhimurium and control sera from non-infected animals. Finally, human antibody fragments (scFv) against these five antigens were selected using antibody phage display and characterised.


CONCLUSIONS
In this work, we identified novel immunogenic proteins of Salmonella Typhimurium and generated antibody fragments against these antigens completely based on phage display. Five immunogenic proteins were validated using a panel of positive and negative sera for prospective applications in diagnostics of Salmonela Typhimurium.
Institut für Biochemie, Biotechnologie und Bioinformatik, Abteilung Biotechnologie, Braunschweig, Germany.
Torsten
T
Meyer
Thomas
T
Schirrmann
André
A
Frenzel
Sebastian
S
Miethe
Janin
J
Stratmann-Selke
Gerald F
GF
Gerlach
Katrin
K
Strutzberg-Minder
Stefan
S
Dübel
Michael
M
Hust
D000818:Animals; D000907:Antibodies, Bacterial / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D000937:Antigen-Antibody Reactions; D001426:Bacterial Proteins / Q000032:analysis* / Q000276:immunology; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D016680:Genome, Bacterial; D006801:Humans; D009842:Oligopeptides / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D016366:Open Reading Frames / Q000235:genetics; D019151:Peptide Library; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012480:Salmonella Infections / Q000175:diagnosis; D012481:Salmonella Infections, Animal / Q000276:immunology / Q000378:metabolism; D012486:Salmonella typhimurium / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013552:Swine
D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D009842:Oligopeptides; D019151:Peptide Library; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies