Cole G T, Kruse D, Seshan K R
Department of Botany, University of Texas, Austin 78713-7640.
Infect Immun. 1991 Jul;59(7):2434-46. doi: 10.1128/iai.59.7.2434-2446.1991.
The occurrence in patients of elevated levels of immunoglobulin M (IgM) precipitin antibody to Coccidioides immitis antigens, which are commonly detected by the immunodiffusion-tube precipitin (TP) assay, is suggestive of primary nondisseminating coccidioidomycosis. We previously demonstrated that the concanavalin A-bound mycelial culture filtrate plus lysate preparation is a source of at least two TP antibody-reactive antigens (TP-Ags), which were identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis as 120- and 110-kDa fractions. Evidence is presented here that the crude filtrate plus lysate preparation contains additional lectin-bound, TP antibody-reactive fractions as well as a component which elicits a complement fixation antibody response in patients. The 120- and 110-kDa fractions were isolated from the antigen complex and further characterized in this paper. Both TP-Ags are glycoproteins and have been shown by immunoelectron microscopy to be colocalized within cytoplasmic vesicles and the wall of spherules. Deglycosylation of these TP-Ags by sodium periodate treatment resulted in a loss in patients of 82 to 95% of IgM adsorption to the antigens as detected by the enzyme-linked immunosorbent assay (ELISA). Comparison of their carbohydrate compositions revealed that mannose and glucose are the predominant monosaccharides of both TP-Ags but only the 120-kDa fraction contained 3-O-methylmannose, a sugar which appears to be unique to C. immitis among the systemic fungal pathogens. We previously showed that 3-O-methylmannose is at least partly responsible for the reactivity of IgM antibody with the 120-kDa TP-Ag. Good correlation was shown between results of immunodiffusion-TP assays and ELISAs of IgM response to both the 120- and 110-kDa fractions by using 70 serum samples from patients with proved coccidioidomycosis. However, only 2.8% (3 of 109) of the serum samples from patients with other mycoses and nonmycotic infections showed IgM adsorption to the 120-kDa TP-Ag as detected by the ELISA, while 21.1% (23 of 109) showed IgM adsorption to the 110-kDa TP-Ag. The 120-kDa TP-Ag is a potentially valuable serodiagnostic reagent for detection of specific IgM by ELISA in patients with primary coccidioidomycosis.
通过免疫扩散管沉淀（TP）试验通常可检测到，患者体内针对粗球孢子菌抗原的免疫球蛋白M（IgM）沉淀抗体水平升高，提示为原发性非播散性球孢子菌病。我们之前证明，伴刀豆球蛋白A结合的菌丝体培养滤液加裂解物制剂是至少两种TP抗体反应性抗原（TP-Ags）的来源，经十二烷基硫酸钠-聚丙烯酰胺凝胶电泳鉴定，这两种抗原为120 kDa和110 kDa的组分。本文提供的证据表明，粗滤液加裂解物制剂还含有其他凝集素结合的、TP抗体反应性组分，以及一种能在患者体内引发补体结合抗体反应的成分。本文从抗原复合物中分离出120 kDa和110 kDa的组分并对其进行了进一步表征。两种TP-Ags均为糖蛋白，免疫电子显微镜显示它们共定位于细胞质囊泡和球形体壁内。经高碘酸钠处理使这些TP-Ags去糖基化后，通过酶联免疫吸附测定（ELISA）检测发现，患者体内IgM对抗原的吸附损失了82%至95%。对它们碳水化合物组成的比较显示，甘露糖和葡萄糖是两种TP-Ags的主要单糖，但只有120 kDa的组分含有3-O-甲基甘露糖，在系统性真菌病原体中，这种糖似乎是粗球孢子菌所特有的。我们之前表明，3-O-甲基甘露糖至少部分负责IgM抗体与120 kDa TP-Ag的反应性。通过对70份已确诊球孢子菌病患者的血清样本进行检测，免疫扩散-TP试验结果与IgM对120 kDa和110 kDa组分反应的ELISA结果之间显示出良好的相关性。然而，在其他真菌病和非真菌性感染患者的血清样本中，只有2.8%（109份中的3份）的样本经ELISA检测显示IgM吸附到120 kDa的TP-Ag上，而21.1%（109份中的23份）的样本显示IgM吸附到110 kDa的TP-Ag上。120 kDa的TP-Ag是一种潜在有价值的血清学诊断试剂，可通过ELISA检测原发性球孢子菌病患者体内的特异性IgM。
