Dasch G A, Samms J R, Williams J C
Infect Immun. 1981 Jan;31(1):276-88. doi: 10.1128/iai.31.1.276-288.1981.
Species-specific antigens from Rickettsia typhi and Rickettsia prowazekii were readily solubilized by French pressure cell extraction or sonication of Renografin density gradient-purified rickettsiae and were identified by rocket immunoelectrophoresis. As measured by quantitative rocket immunoelectrophoresis, the species-specific typhus rocket antigens (STRAs) appeared to be proteins; they were denatured by heating at 56 degrees C for 30 min but not by 50 degrees C treatment, and they were sensitive to pronase and trypsin but were not affected by periodate oxidation, glycosidases of various specificities, phospholipase A, or lipase. STRAs from both R. typhi and R. prowazekii were separated from common antigens by DE52 column chromatography of 100,000-X-g supernatant fractions of rickettsial extracts. The purified STRAs were characterized by crossed immunoelectrophoresis, by polyacrylamide gel electrophoresis on Davis and sodium dodecyl sulfate gels, and by an enzyme-linked immunosorbent assay. The two purified STRAs were proteins with similar native electrophoretic mobilities in agarose and polyacrylamide gels, and these proteins had similar polypeptide patterns on sodium dodecyl sulfate gels. Most of the STRA activity migrated as a single protein band on sodium dodecyl sulfate-polyacrylamide and Davis polyacrylamide gels, although minor protein bands with STRA activity were also detected. The major STRA proteins constituted 10 to 15% of the total cellular protein of R. typhi and R. prowazekii. According to sensitive enzyme-linked immunosorbent assay titrations, the STRA of R. prowazekii had substantial cross-reactivity with rabbit antiserum prepared against R. typhi, as shown also by rocket immunoelectrophoresis, whereas the STRA of R. typhi reacted only very weakly with antiserum prepared against R. prowazekii according to the enzyme-linked immunosorbent assay and not at all according to rocket immunoelectrophoresis.
通过对泛影葡胺密度梯度纯化的立克次氏体进行法国压力细胞提取或超声处理，可轻松溶解来自伤寒立克次氏体和普氏立克次氏体的种特异性抗原，并通过火箭免疫电泳进行鉴定。通过定量火箭免疫电泳测量，种特异性斑疹伤寒火箭抗原（STRA）似乎是蛋白质；它们在56℃加热30分钟会变性，但在50℃处理时不会变性，并且它们对链霉蛋白酶和胰蛋白酶敏感，但不受高碘酸盐氧化、各种特异性糖苷酶、磷脂酶A或脂肪酶的影响。通过对立克次氏体提取物100,000-X-g上清液部分进行DE52柱色谱，可将伤寒立克次氏体和普氏立克次氏体的STRA与共同抗原分离。纯化的STRA通过交叉免疫电泳、在戴维斯凝胶和十二烷基硫酸钠凝胶上进行聚丙烯酰胺凝胶电泳以及酶联免疫吸附测定进行表征。两种纯化的STRA是在琼脂糖和聚丙烯酰胺凝胶中具有相似天然电泳迁移率的蛋白质，并且这些蛋白质在十二烷基硫酸钠凝胶上具有相似的多肽模式。尽管也检测到了具有STRA活性的次要蛋白带，但在十二烷基硫酸钠-聚丙烯酰胺和戴维斯聚丙烯酰胺凝胶上，大部分STRA活性以单一蛋白带形式迁移。主要的STRA蛋白占伤寒立克次氏体和普氏立克次氏体总细胞蛋白的10%至15%。根据灵敏的酶联免疫吸附测定滴定，普氏立克次氏体的STRA与针对伤寒立克次氏体制备的兔抗血清具有显著的交叉反应性，火箭免疫电泳也显示了这一点，而根据酶联免疫吸附测定，伤寒立克次氏体的STRA与针对普氏立克次氏体制备的抗血清反应非常弱，根据火箭免疫电泳则根本不反应。
