Use of a recombinant 170-kilodalton surface antigen of Entamoeba histolytica for serodiagnosis of amebiasis and identification of immunodominant domains of the native molecule.
We expressed the gene that encodes one of the major surface antigens of Entamoeba histolytica, the 170-kDa protein (1,270 amino acids), as a glutathione S-transferase fusion protein containing amino acids 1 to 1202 (lacking the putative transmembrane and cytoplasmic regions) and as separate fusion proteins containing each of three major domains of the 170-kDa molecule. Lysates from bacteria induced to express one of these proteins were used as the target antigens in a Western blot (immunoblot) analysis to determine whether a recombinant 170-kDa antigen could serve as the basis for a serologic test used to detect invasive amebiasis and whether there are differences in humoral immunogenicity among the three major domains of the 170-kDa antigen. Among patients with invasive amebiasis from three major areas where the disease is endemic and two sites in the United States, 54 (90%) of 60 had antibodies to the recombinant 170-kDa protein. Among 37 patients from regions where the disease is endemic and 20 patients from the United States without amebic disease, 1 (2%) of 57 had antibodies to the recombinant 170-kDa protein. We found significant differences in seroreactivity to each of three major domains of the molecule among patients seropositive for the complete construct, ranging from 100% seroreactivity with the fusion protein containing the domain designated cysteine rich and 89% seropositivity with the fusion protein incorporating a portion of the region designated cysteine poor to only 9% seropositivity for the fusion protein containing the pseudorepeat domain. Our study indicates that a serologic test based on the recombinant 170-kDA antigen could serve as a highly sensitive and specific test for acute invasive amebiasis.
Zhang
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Y
Department of Medicine, Washington University School of Medicine, St. Louis, Missouri 63110.
Li
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E
Jackson
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TF
Zhang
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T
Gathiram
V
V
Stanley
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SL
D000818:Animals; D000953:Antigens, Protozoan / Q000235:genetics; D000954:Antigens, Surface / Q000235:genetics; D015153:Blotting, Western / Q000379:methods / Q000706:statistics & numerical data; D016054:DNA, Protozoan / Q000235:genetics; D004748:Entamoeba histolytica / Q000235:genetics / Q000276:immunology*; D004749:Entamoebiasis / Q000175:diagnosis*; D005069:Evaluation Studies as Topic; D006801:Humans; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012680:Sensitivity and Specificity; D012698:Serologic Tests / Q000379:methods / Q000706:statistics & numerical data
D000953:Antigens, Protozoan; D000954:Antigens, Surface; D016054:DNA, Protozoan; D011994:Recombinant Proteins