摘要
目的 研究嗜水气单胞菌hec毒素对红细胞的作用 ,为阵发性睡眠性血红蛋白尿症(PNH)的临床诊断提供一种新方法。方法 从患暴发性传染病的家养鲫鱼分离到嗜水气单胞菌 ,其培养上清经硫酸铵沉淀获得粗提毒素 ,经DEAE纤维素层析得提纯毒素。粗提毒素作用于PNH和其他贫血患者及正常人的红细胞 ,采用 96孔板在波长 6 30nm酶标仪比色 ;同时用CD59单抗和FITC标记的羊抗鼠IgG标记红细胞 ,以流式细胞仪分析毒素作用前后CD59标记率。结果 对 15例PNH患者、15名正常人、15例非PNH贫血患者的红细胞 ,进行hec毒素试验 ,发现毒素在 0 .12 5g/L、37℃作用 2 0min后 ,正常人和非PNH贫血患者的红细胞完全溶解 ,而PNH患者存留红细胞的百分率与CD59标记率成反比。毒素作用前后流式细胞仪检测结果表明 ,毒素可以特异地作用于CD59阳性细胞。结论 PNH患者的红细胞比其他非PNH贫血患者及正常人的红细胞有明显的抗毒素溶破作用 ,且毒素作用后存留细胞的百分数反映CD59标记率 ,故可作为PNH诊断的敏感、特异、简便可行的新诊断方法。
Objective To study the feasibility of diagnosing paroxysmal nocturnal hemoglobinuria(PNH) with hec toxin (secreted by Aeromonus hydrophila) test. Methods The crude hec toxin was extracted from the culture medium of Aeromonus hydrophila by precipitating with saturated (NH 4) 2SO 4 and then purified through DEAE52 . This crude hec toxin was used to act on red blood cells(RBCs) from patients with PNH, non-PNH anemias, and normal persons. Absorbance at 630nm was measured to quantitate the extent of hemolysis. Hec treated and untreated RBCs were both stained with anti CD 59 monoclonal antibody and FITC labelled goat-anti-mouse IgG. The percentage of CD 59 + cells was detected by flow cytometry. Results After hec toxin treatment, RBCs from PNH patients showed resistance to the toxin hemolysis, which was negatively related to the CD 59 + cells percentage, while RBCs from non-PNH were lysed totally. Conclusion RBCs from PNH have obvious resistance to the hemolytic effect of hec toxin, and the percentage of remained unhemolytic cells reflect the severity of PNH. Resistance detection of RBCs to hec toxin can be used for the diagnosis of PNH.
出处
《中华血液学杂志》
CSCD
北大核心
2000年第10期517-520,共4页
Chinese Journal of Hematology
基金
国家自然科学基金!资助项目 (39779334)
卫生部基金!资助项目 (98 1 0 2 5 )
北京市自然科学基金!资助项目 (7982 0 32 )
关键词
阵发性睡眠性血红蛋白尿
嗜水气单胞菌
hec毒素
Hemoglobinuria
paroxysmal
hec toxin
Antigen
CD
59
Glycosylphosphatidylinositol protein
Aeromonus hydrophila
