RESUMO
In Fasciola parasites fatty acid binding proteins (FABPs) are the carrier proteins that help in the uptake of fatty acids from the hosts' fluids. Attempts have been made to utilize both native and recombinant FABP (rFABP) for immunodiagnosis and vaccine development for fasciolosis. In this study, we have produced a number of monoclonal antibodies (MoAbs) against rFABP of Fasciola gigantica. These MoAbs were initially screened against rFABP by ELISA and then tested for their specificities by immunoblotting. Five stable clones were selected and characterized further: four of them were of the isotype IgG(1) while one clone was IgG(2a). All the MoAbs reacted with rFABP which has a molecular weight (MW) of 20 kD and with at least two isoforms of native proteins at MW 14.5 kD that were present in the tegumental antigen (TA) and crude worm extracts, and the excretion-secretion materials. Immunoperoxidase staining of frozen sections of adult parasites by using these MoAbs as primary antibodies indicated that FABP were present in high concentration in the parenchymal cells and reproductive tissues, in low concentration in the tegument and caecal epithelium. All MoAbs cross-reacted with a 14.5 kD antigen present in the whole body (WB) extract of Schistosoma mansoni, while no cross-reactivities were detected with antigens from Eurytrema pancreaticum and Paramphistomum spp.
Assuntos
Anticorpos Anti-Helmínticos/imunologia , Anticorpos Monoclonais/imunologia , Proteínas de Transporte/imunologia , Fasciola/imunologia , Fasciolíase/veterinária , Proteínas de Neoplasias , Animais , Especificidade de Anticorpos , Proteínas de Transporte/isolamento & purificação , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática/veterinária , Fasciolíase/diagnóstico , Proteínas de Ligação a Ácido Graxo , Proteínas de Helminto/imunologia , Proteínas de Helminto/isolamento & purificação , Immunoblotting/veterinária , Técnicas Imunoenzimáticas/veterinária , Imunoglobulina G/imunologia , Peso Molecular , Isoformas de Proteínas , Proteínas Recombinantes/imunologiaRESUMO
The genomic organization of the knob protein (KP) gene of knobby (K+) and knobless (K-) variants of the Thai isolate NT 108 and the Gambian isolate FCR-3 are compared. The restriction enzyme maps and the chromosomal location of the KP gene of K+ variants of both isolates are apparently identical. A comparison of the susceptibility of the KP gene to deletions and the extent of deletion of chromosomal DNA in K- variants of each geographical isolate suggests isolate-specificity of the stability of the deleted DNA sequences. With the exception of a mutant K- population of FCR-3, which could not be distinguished from K+ FCR-3, in all other K- variants of both geographical isolates the deletion of the KP gene was accompanied by the loss of several hundred base pairs of DNA from chromosome 2. The deletion resulted in the localization of the mutant KP gene in proximity to telomeric DNA sequences.
Assuntos
Proteínas de Membrana/genética , Peptídeos/genética , Plasmodium falciparum/genética , Animais , Southern Blotting , DNA/genética , Sondas de DNA , Desoxirribonuclease HindIII , Eletroforese em Gel de Ágar , Gâmbia , Microscopia Eletrônica , Hibridização de Ácido Nucleico , Plasmodium falciparum/ultraestrutura , Proteínas de Protozoários , Mapeamento por Restrição , TailândiaRESUMO
A cDNA library constructed from ring-stage RNA isolated from Plasmodium falciparum FCR-3/Gambia was screened with immune human serum and two related positive clones were isolated. Nucleotide sequence analysis of these recombinant clones revealed an open translational reading frame for 681 amino acids with a calculated molecular weight of 74.3 kDa. The deduced amino acid sequence of the polypeptide shows extensive homology to several heat shock proteins (hsp) which have been described. Northern and Southern hybridization analysis indicates that P. falciparum has a second gene which shares common sequences with the hsp gene described in this study.