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| 題 名 | 宿主細胞(RK13細胞)膜上弓蟲分泌性抗原接受器之研究 |
|---|---|
| 作 者 | 張甘楠; | 書刊名 | 臺灣畜牧獸醫學會會報 |
| 卷 期 | 65:3 1995.09[民84.09] |
| 頁 次 | 頁255-264 |
| 分類號 | 437.21 |
| 關鍵詞 | 穿透性加強因子; 分泌性抗原; 弓蟲; Penetration-enchancing factor; Secretory antigen; Toxoplasma gondii; |
| 語 文 | 中文(Chinese) |
| 中文摘要 | 以增殖型弓蟲(Rh株)接種於RK13細胞4小時後,至所有弓蟲均侵入RK13細胞內,取該細胞培養液Cell Free之上層液可莘取弓蟲分泌抗原,該抗原是弓蟲對宿主細胞(RK13細胞)之穿透加強因子(Secretory Penetration-enhancing Factor,SPEF),以此SPEF 25μg/well 加入RK13單層胞經37℃,4小時後,以106增殖型弓蟲接種,接種後18小時與不加SPEF的對照組相比較,評估試驗組之感染單位相對數(relative number of infective units,RNIU)與對照繆之感染單位相對數相互比較結果其比為1.56:1;且72小時後接種增殖型弓蟲(106/well)的試驗組比對照組呈現較單重的CPE(cytopathic effect),試驗組之弓蟲破壞RK13細胞產生CPE之百分比約(25%)多於對照組(約5%),顯示穿透加強因子確能增強弓蟲對RK13細胞之穿透及感染能力。進一步採用Toxoplasma gondii所研製之單株抗體(McAb/TGCM,IgG1, K)來檢測,可證明單株體可完全阻斷增殖型弓蟲(Rh株)侵入RK13細胞內,以SPEF為橋樑介於RK13細胞與單株抗體之間行間接螢光抗體法結果顯示RK13細胞有強烈的特異性螢光反應,證實RK13細胞膜上具有弓蟲SPEF的接受器存在,此亦顯示此SPEF為弓蟲對RK13細胞的一種吸附性的蛋白。以西方轉漬法分析結果獲知弓蟲對RK13細胞之皆泌性穿透加強因子其主要成分是弓蟲分泌性抗原中的胜□76KD。 |
| 英文摘要 | A secretory penetration-enhancing factor (SPEF) was extracted from the cell-free supernatant of RK13 cell cultures inoculated with tachyzoites of Toxoplasma gondii (Rh strain) at 37℃ for 4 hours after all the tachyzoites had penetrated into the cell. To elucidate the receptors for SPEF of Toxoplasma gondii, the monolayer of RK13 cells in 8-well chamber slide were pretreated with 25 wg of SPEF at 37℃ for 4 hours, followed by inoculation with 106 tachyzoites per well and incubated at 37℃ for 18 hours. The control group was inoculated with 106 tachyzoites per well without SPEF pretreatment. The penetration-enhancing activity as expressed by the ratio of the relative number of infective units (RNIU) with SPEF pretreatment to those of without SPEF treatment, was approximately 1 to 1.56 times in terms of penetration enhancement of T. gondii into the host cells (RK13). Moreover, seventy-two hours after inoculation, the RK13 monolayer cells with SPEF pretreatment showed more CPE (cytopathic effect) than those of the control group. However, a monoclonal antibody (McAb/TGCM, IgG1, K) derived from T.gondii tachyzoites could complete block the penetration of tachyzoites into the RK13 cells. The McAb/TGCM was used to check for the existence of receptors for SPEF on the cell membrane of RK13 cells by using indirect fluorescent antibody (IFA) test. It was demonstrated that the RK13 cells showed strongly specific reaction in IFA test when SPEF of T. fonidii was added to crosslink the RK13 cells. Thus, it is suggested that eceptors for SPEF are present on the cells. The SPEF were also shown to be polypeptides with molecular weight of 76KD when analyzed by western blotting assay. |
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