RESUMO
Monomeric and chimeric synthetic peptides were used as coating antigens in four different mixtures in a solid phase immunoassay to select an optimal combination for the detection of antibodies to human T-cell lymphotropic virus (HTLV) in serum samples. The peptides, P-13 (gp21 I), Q5 (gp21 II)-GG-(gp46 II), and Q (gp46 I)-GG-(p19 I), represent immunodominant sequences from transmembrane protein (gp21), envelope protein (gp46), and core protein (p19) of HTLV I/II viruses; they were the most antigenic and specific peptides in previous studies. The sequences of the chimeric peptides were separated by two glycine residues. An indirect UltramicroEnzyme-linked immunosorbent assay (UMELISA) was used to evaluate the antigenicity of these peptide mixtures by using samples from anti-HTLV I/II PRP205(M), (n = 20), HTLV I-infected individuals from Cuba (n = 7), and HTLV I-positive sera from Colombia and Chile (n = 9). The specificity was evaluated with healthy blood donor sera (n = 300), anti-HIV 1-positive samples (n = 10), and other seropositive samples to different infectious agents. The highest sensitivity and specificity was obtained with mixture 1, which could be very useful in the immunodiagnostic of HTLV infection.
Assuntos
Anticorpos Anti-HTLV-I/imunologia , Antígenos HTLV-I/imunologia , Infecções por HTLV-I/diagnóstico , Infecções por HTLV-I/imunologia , Anticorpos Anti-HTLV-II/imunologia , Antígenos HTLV-II/imunologia , Infecções por HTLV-II/imunologia , Peptídeos/imunologia , Ensaio de Imunoadsorção Enzimática/métodos , Anticorpos Anti-HTLV-I/sangue , Antígenos HTLV-I/química , Infecções por HTLV-I/sangue , Anticorpos Anti-HTLV-II/sangue , Antígenos HTLV-II/química , Infecções por HTLV-II/sangue , Humanos , Peptídeos/química , Sensibilidade e EspecificidadeRESUMO
A chimeric synthetic peptide incorporating immunodominant epitope of the p19 gag protein (116-134) and the gp46 env protein (178-200) of HTLV-II virus, separated by two glycine residues, was synthesized by conventional solid-phase peptide synthesis. The antigenic activity of this peptide was evaluated by Ultramicro Enzyme-linked immunosorbent assay (UMELISA) by using panels of anti-HTLV-II positive sera (n = 9), anti-HTLV-I/II positive sera (n = 2), HTLV-positive (untypeable) serum samples (n = 1),and anti-HTLV-I positive sera (n = 14), while specificity was evaluated with samples from healthy blood donors (n = 20). The efficacy of the chimeric peptide in solid-phase immunoassays was compared with the monomeric peptides. Data demonstrated that the chimeric peptide was the most reactive because it detected antibodies to virus efficiently. This may be related to peptide adsorption to the solid surface and epitope accessibility to the antibodies. The results indicate that chimeric peptide as coating antigen is very useful for the immunodiagnosis of HTLV-II infection.
Assuntos
Produtos do Gene env/química , Antígenos HTLV-II/química , Antígenos HTLV-II/imunologia , Leucemia de Células T/sangue , Leucemia de Células T/imunologia , Proteínas dos Microtúbulos , Fosfoproteínas/química , Proteínas Oncogênicas de Retroviridae/química , Produtos do Gene env/imunologia , Humanos , Leucemia de Células T/diagnóstico , Peptídeos/síntese química , Peptídeos/imunologia , Fosfoproteínas/imunologia , Proteínas Recombinantes de Fusão/síntese química , Proteínas Recombinantes de Fusão/imunologia , Proteínas Oncogênicas de Retroviridae/imunologia , Estatmina , Produtos do Gene env do Vírus da Imunodeficiência HumanaRESUMO
Two chimeric synthetic peptides incorporating immunodominant sequences from HTLV-II virus were synthesized. Monomeric peptides P2 and P3 represent sequences from transmembrane protein (gp21) and envelope protein (gp46) of the virus. The peptide P2 is a gp21 (370-396) sequence and the peptide P3 is a gp46 (178-205) sequence. Those peptides were arranged in a way that permits one to obtain different combinations of chimeric peptides (P2-GG-P3 and P3-GG-P2), separated by two glycine residues as spacer arms. The antigenic activity of these peptides was evaluated by UltramicroEnzyme-linked immunosorbent assay (UMELISA) by using panels anti-HTLV-II-positive sera (n = 11), anti-HTLV-I/II-positive sera (n = 2), HTLV-positive (untypeable) serum samples (n = 2), and anti-HTLV-I-positive sera (n = 22), while specificity was evaluated with anti-HIV-positive samples (n = 19) and samples from healthy blood donors (n = 30). The efficacy of the chimeric peptides in solid-phase immunoassays was compared with the monomeric peptides and a mixture of the monomeric peptides. Higher sensitivity was observed for chimeric peptide Q5 assay. Those results may be related to a higher peptide adsorption capacity to the solid surface and for epitope accessibility to the antibodies. This chimeric peptide would be very useful for HTLV-II diagnostic.
Assuntos
Produtos do Gene env/imunologia , Anticorpos Anti-HTLV-II/sangue , Antígenos HTLV-II/química , Vírus Linfotrópico T Tipo 2 Humano/imunologia , Proteínas Oncogênicas de Retroviridae/imunologia , Sequência de Aminoácidos , Ensaio de Imunoadsorção Enzimática , Produtos do Gene env/química , Produtos do Gene env/genética , Antígenos HTLV-II/genética , Infecções por HTLV-II/diagnóstico , Infecções por HTLV-II/imunologia , Vírus Linfotrópico T Tipo 2 Humano/genética , Humanos , Epitopos Imunodominantes/química , Epitopos Imunodominantes/genética , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Proteínas Oncogênicas de Retroviridae/química , Proteínas Oncogênicas de Retroviridae/genética , Produtos do Gene env do Vírus da Imunodeficiência HumanaRESUMO
A recombinant protein of the human T cell lymphotropic virus type I (HTLV-I) gp46 outer membrane envelope, MTA-4 (residues 129-203), reacted by Western blot with sera from HTLV-I-infected individuals from the United States and Jamaica but not with 24 (10%) of 242 Japanese sera. A related gp46 recombinant protein, MTA-1 (residues 162-209), reacted with all 58 sera from HTLV-I-infected US and Jamaican individuals and 238 of 242 sera from infected Japanese (combined sensitivity of 99%). Neither recombinant showed reactivity to sera from HTLV-II-infected individuals or uninfected controls. The reactivity of recombinant proteins containing the region of HTLV-II gp46 analogous to MTA-1 was also evaluated by Western blot: GH2-K15 (residues 157-205) and GH2-K55 (residues 162-205) reacted with 88 (98%) and 89 (99%), respectively, of 90 sera from HTLV-II-infected individuals but not with sera from HTLV-I-infected individuals or uninfected controls. These recombinant proteins should permit the development of assays to unambiguously confirm and differentiate HTLV-I and HTLV-II infections.