Production and characterization of a monoclonal antibody against recombinant glutathione S-transferase (GST) of Fasciola gigantica.
A monoclonal antibody (MoAb) against a recombinant glutathione S-transferase (rGST) of F. gigantica was produced in BALB/c mice. Reactivity and specificity of this monoclonal antibody was assessed by ELISA and immunoblotting. Six stable clones, namely 3A3, 3B2, 3C6, 4A6, 4B1 and 4D6 were obtained, All these MoAb reacted with rGST and native GST at a molecular weight of 28 kDa and found to be IgG1, kappa-light chain isotypes. These MoAb cross-reacted with Schistosoma mansoni and Schistosoma japonicum antigens at molecular weights of 28 and 26 kDa, respectively, but no cross-reactions were detected with antigens of Eurytrema and Paramphistomum spp. The localization of GST in metacercaria, 7-week-old juvenile and adult F. gigantica was performed by immunofluorescence technique, using MoAb as well as polyclonal antibody (PoAb) to the native protein as probes. In general, all clones of MoAb gave similar results and the pattern was quite similar to staining by PoAb. The fluorescence was intense, which implied the presence of a high concentration of GST in the parenchymal tissue in all stages of the parasite. However, the parenchymal cells were not evenly stained which implied the existence of subpopulations of this cell type with regard to GST production and storage. In addition, in adult and juvenile stages a moderate fluorescence was present in the basal layer of the tegument, while light fluorescence was observed in the caecal epithelium, cells in the ovary, testis and vitelline gland of the adult. In the metacercaria stage, in addition to parenchymal tissue, the tegument and tegumental cells were stained relatively more intense with MoAb and PoAb than in other stages.
Khawsuk
Witoon
W
Department of Anatomy, Faculty of Science, Mahidol University, Bangkok, Thailand.
Soonklang
Nantawan
N
Grams
Rudi
R
Vichasri-Grams
Suksiri
S
Wanichanon
Chaitip
C
Meepool
Ardool
A
Chaithirayanon
Kulathida
K
Ardseungneon
Pissanee
P
Viyanant
Vithoon
V
Upathum
Suchart Edward
SE
Sobhon
Prasert
P
D000818:Animals; D000909:Antibodies, Helminth; D000911:Antibodies, Monoclonal / Q000737:chemistry; D000918:Antibody Specificity; D000947:Antigens, Helminth / Q000276:immunology*; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005209:Fasciola / Q000276:immunology*; D005455:Fluorescent Antibody Technique; D005982:Glutathione Transferase / Q000276:immunology*; D015151:Immunoblotting; D008018:Life Cycle Stages / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D011994:Recombinant Proteins / Q000276:immunology; D012680:Sensitivity and Specificity
D000909:Antibodies, Helminth; D000911:Antibodies, Monoclonal; D000947:Antigens, Helminth; D011994:Recombinant Proteins; D005982:Glutathione Transferase