Isolation of T-cell antigens by using a recombinant protein library and its application to the identification of novel vaccine candidates against schistosomiasis.
We present here a novel approach to identify T-cell antigens from any infectious agent by use of a library of purified recombinant proteins. Essential features of this strategy include (i) a highly efficient cDNA cloning system which negatively selects against nonrecombinant transformants by making use of the bacterial EcoK restriction system, (ii) affinity staining of cDNA clones expressing recombinant proteins, and (iii) a procedure of simultaneous purification of recombinant proteins from large numbers of isolated clones (representing the protein library) in a single step from pools consisting of up to 24 individual clones. The feasibility of the screening system was confirmed by constructing a protein library of the human parasite Schistosoma mansoni. The recombinant antigens of this library were used to stimulate CD4(+) T cells derived from the axillary lymph nodes of mice vaccinated with irradiated cercariae. In initial screening experiments, we detected parasite-specific proliferation and gamma interferon (IFN-gamma) secretion in response to several pools of cDNA clones. Further analysis of one particular pool revealed that only one of its constituents stimulated considerable IFN-gamma secretion by CD4(+) T cells and that the expressed antigen is identical to a small fragment of myosin heavy chain.
Eberl
M
M
Biochemisches Institut, Justus-Liebig-Universität Giessen, 35392 Giessen, Germany. me10@york.ac.uk
Mountford
A P
AP
Jankovic
D
D
Beck
E
E
D000818:Animals; D017951:Antigen Presentation; D000947:Antigens, Helminth / Q000276:immunology*; D003602:Cytotoxicity, Immunologic; D006801:Humans; D051379:Mice; D019151:Peptide Library; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012550:Schistosoma mansoni / Q000276:immunology*; D012555:Schistosomiasis mansoni / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology*; D014612:Vaccines / Q000235:genetics / Q000276:immunology*
D000947:Antigens, Helminth; D019151:Peptide Library; D011948:Receptors, Antigen, T-Cell; D011994:Recombinant Proteins; D014612:Vaccines