| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2003: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 2002: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Research Abstract |
We have recently identified a 150-kDa surface antigen of Entamoeba histolytica as an intermediate subunit (Igl) of galactose-and N-acetyl-D-galactosamine-inhibitable lectin. In the present study, full-length Igl except for the signal sequences [amino acid (aa) 14-1,088] and three fragments of Igl, the N-terminal part (aa 14-382), middle part (aa 294-753), and C-terminal part (aa 603-1,088), were prepared in Escherichia coli and the reactivity of these recombinant proteins with sera from patients with amebiasis was examined by means of enzyme-linked immunosorbent assay (ELISA). Sera from symptomatic patients with amebic liver abscess or amebic colitis, sera from asymptomatic cyst passers, sera from individuals with other protozoan infections, and sera from healthy controls were used. Sensitivity and specificity of the recombinant full-length Igl in the ELISA were 90% and 94% respectively. When three fragments were used as antigens in the ELISA, sensitivities were 56% in the N-terminus,
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92% in the middle part, and 97% in the C-terminus. Specificities of the three antigens were 96% in the N-terminus and 99% in both the middle and C-terminus fragments. None of the sera from asymptomatic cyst passers was negative when F-Igl, M-Igl and C-Igl were used as the antigen. On the other hand, in the cases of ELISA using M-Igl and C-Igl, false-negative results were detected in only the colitis cases. These observations suggest that the antibodies which recognized the epitopes located in M-Igl and C-Jgl may function to prevent the invasion of trophozoites into host tissues and that the carboxyl terminus of Igl is an especially useful antigen for the serodiagnosis of amebiasis. When the reactivity of adherence-inhibitory monoclonal antibodies to various recombinant fragments was also examined, one of the epitope was localized in the part of aa 989-1, 088. Liver abscess formation of hamsters was significantly inhibited by the immunization with native Igl, but not with the F-Igl, suggesting that the epitope (s) on sugar chain(s) of Igi may also be important for the induction of protective immunity. Two Igl genes, coding 1, 110 and 1, 106 aa, were also cloned from Entamoeba dispar and were further analyzed. Less
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