Legionella pneumophila surface antigens cloned and expressed in Escherichia coli are translocated to the host cell surface and interact with specific anti-Legionella antibodies.
Escherichia coli clones that express Legionella pneumophila antigens were isolated from a plasmid genomic library, and their antigens were characterized by immunoblotting with rabbit anti-L. pneumophila sera. Because previous studies of L. pneumophila antigens by whole-cell radioimmunoprecipitation suggested that comigrating native antigens were surface localized, we conducted experiments to determine if the cloned antigens were surface expressed in E. coli. Aliquots of antisera were absorbed by intact cells of three representative antigen-producing E. coli clones, and surface-bound antibodies were acid eluted from the intact cells. Immunoblots made with selectively absorbed antisera and eluted antibodies confirmed that reactivity to the homologous cloned antigens could be specifically absorbed from the antisera and then eluted from the cells, demonstrating a surface (antibody-accessible) localization in the cloned state. Antibodies eluted from the surface of an E. coli clone that expressed a 19-kilodalton antigen reacted with the surface of L. pneumophila in a liquid-phase, whole-cell enzyme-linked immunosorbent assay. In addition, intact cells of this clone were used in an enzyme-linked immunosorbent assay to detect serum antibody. E. coli cells that express foreign antigens on their surfaces can be used to develop antigen-specific immunoassays and to affinity purify monospecific antibodies.
D000818:Animals; D000936:Antigen-Antibody Complex; D000942:Antigens, Bacterial / Q000032:analysis / Q000235:genetics*; D000954:Antigens, Surface / Q000032:analysis / Q000235:genetics*; D003001:Cloning, Molecular; D006224:Cricetinae; D004926:Escherichia coli / Q000235:genetics*; D007875:Legionella / Q000235:genetics / Q000276:immunology*; D008970:Molecular Weight; D014178:Translocation, Genetic
D000936:Antigen-Antibody Complex; D000942:Antigens, Bacterial; D000954:Antigens, Surface