title: Production of antigen-specific human IgGs by in vitro immunization.
abstract: BACKGROUND
  We previously developed in vitro immunization based on a fusion protein containing the transcriptional transactivator (Tat) of human immunodeficiency virus and a double domain, called ZZ, derived from protein A of Staphylococcus aureus. In this approach, naïve human peripheral blood mononuclear cells (PBMCs) trigger a specific IgM antibody (Ab) response in the presence of ZZTat. In the present study, we attempted to raise a specific IgG Ab response.
  
  
  RESULTS
  We found that PBMCs incubated with ZZTat and a mixture containing anti-CD40, IL4 and IL21 secrete anti-Tat IgG Abs in their supernatants, indicating that the cytokine cocktail provides an isotypic switch. Then, we deciphered the Tat determinant involved in the phenomenon and found that it is located in the region 22-57 and that, within this region, the cysteine-rich domain and the basic residues play a crucial role. Finally, we prepared a fusion protein containing a fragment derived from the NY-ESO-1 cancer/testis antigen (Ag) and showed that PBMCs incubated with ZZfNY-ESO-1Tat trigger a specific anti-fNY-ESO-1 IgG Ab response, which demonstrates the possibility of transferring immunizing ability to an Ag unrelated to Tat.
  
  
  CONCLUSIONS
  Our ZZTat-based in vitro immunization approach that offers the possibility to raise an IgG Ab response against NY-ESO-1 might represent a valuable first stage for the generation of fully human IgG specific Abs.
authors.affiliation: University of Paris Diderot, Paris, France. anne.wijkhuisen@cea.fr.|CEA, Institut de Biologie et de Technologie de Saclay (iBiTec-S), Service de Pharmacologie et d'Immunoanalyse (SPI), 91191, Gif sur Yvette, France. anne.wijkhuisen@cea.fr.
authors.forename: A
authors.initials: A
authors.lastname: Wijkhuisen
authors.affiliation: CEA, Institut de Biologie et de Technologie de Saclay (iBiTec-S), Service de Pharmacologie et d'Immunoanalyse (SPI), 91191, Gif sur Yvette, France.
authors.forename: A
authors.initials: A
authors.lastname: Savatier
authors.affiliation: CEA, Institut de Biologie et de Technologie de Saclay (iBiTec-S), Service de Pharmacologie et d'Immunoanalyse (SPI), 91191, Gif sur Yvette, France.
authors.forename: N
authors.initials: N
authors.lastname: Cordeiro
authors.affiliation: CEA, Institut de Biologie et de Technologie de Saclay (iBiTec-S), Service de Pharmacologie et d'Immunoanalyse (SPI), 91191, Gif sur Yvette, France.
authors.forename: M
authors.initials: M
authors.lastname: Léonetti
mesh_terms: D002478:Cells, Cultured; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology* / Q000378:metabolism*; D007963:Leukocytes, Mononuclear / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D013205:Staphylococcal Protein A / Q000235:genetics / Q000276:immunology; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000235:genetics
chemical_list: D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D013205:Staphylococcal Protein A; D054322:tat Gene Products, Human Immunodeficiency Virus