title: Analysis of Leishmania chagasi by 2-D difference gel electrophoresis (2-D DIGE) and immunoproteomic: identification of novel candidate antigens for diagnostic tests and vaccine.
abstract: Identification of novel antigens is essential for developing new diagnostic tests and vaccines. We used DIGE to compare protein expression in amastigote and promastigote forms of Leishmania chagasi. Nine hundred amastigote and promastigote spots were visualized. Five amastigote-specific, 25 promastigote-specific, and 10 proteins shared by the two parasite stages were identified. Furthermore, 41 proteins were identified in the Western blot employing 2-DE and sera from infected dogs. From these proteins, 3 and 38 were reactive with IgM and total IgG, respectively. The proteins recognized by total IgG presented different patterns in terms of their recognition by IgG1 and/or IgG2 isotypes. All the proteins selected by Western blot were mapped for B-cell epitopes. One hundred and eighty peptides were submitted to SPOT synthesis and immunoassay. A total of 25 peptides were shown of interest for serodiagnosis to visceral leishmaniasis. In addition, all proteins identified in this study were mapped for T cell epitopes by using the NetCTL software, and candidates for vaccine development were selected. Therefore, a large-scale screening of L. chagasi proteome was performed to identify new B and T cell epitopes with potential use for developing diagnostic tests and vaccines.
authors.affiliation: Universidade Federal de Minas Gerais, Instituto de Ciências Biológicas, Departamento de Bioquímica e Imunologia, 31270-910 Belo Horizonte, Minas Gerais, Brasil.
authors.forename: Míriam M
authors.initials: MM
authors.lastname: Costa
authors.forename: Hélida M
authors.initials: HM
authors.lastname: Andrade
authors.forename: Daniella C
authors.initials: DC
authors.lastname: Bartholomeu
authors.forename: Leandro M
authors.initials: LM
authors.lastname: Freitas
authors.forename: Simone F
authors.initials: SF
authors.lastname: Pires
authors.forename: Alexander D
authors.initials: AD
authors.lastname: Chapeaurouge
authors.forename: Jonas
authors.initials: J
authors.lastname: Perales
authors.forename: André T
authors.initials: AT
authors.lastname: Ferreira
authors.forename: Mário S
authors.initials: MS
authors.lastname: Giusta
authors.forename: Maria N
authors.initials: MN
authors.lastname: Melo
authors.forename: Ricardo T
authors.initials: RT
authors.lastname: Gazzinelli
mesh_terms: D000818:Animals; D000953:Antigens, Protozoan / Q000276:immunology*; D015153:Blotting, Western; D019295:Computational Biology; D004285:Dogs; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D018985:Epitopes, B-Lymphocyte / Q000235:genetics*; D007118:Immunoassay; D007891:Leishmania / Q000276:immunology*; D007898:Leishmaniasis, Visceral / Q000175:diagnosis*; D040901:Proteomics / Q000379:methods*; D012698:Serologic Tests / Q000379:methods*; D013045:Species Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000953:Antigens, Protozoan; D018985:Epitopes, B-Lymphocyte