摘要
目的探讨甲醛固定、石蜡包埋的肾穿刺组织做免疫荧光染色的可行性。方法选择10例膜性肾病、25例IgA肾病及10例狼疮肾炎患者的肾穿刺组织。对石蜡切片进行脱腊、修复抗原及直接免疫荧光染色,并将染色结果与冰冻切片染色结果进行比较。结果在本实验条件下,二甲苯浸泡30 min、胃蛋白酶消化10 min为最适脱蜡及抗原修复条件,抗体稀释至特异荧光强度足够及非特异荧光背底最弱为当。应用上述条件对石蜡切片进行直接免疫荧光染色,其结果与冰冻切片染色的诊断符合率达100%,而石蜡切片具有组织结构保存好、免疫沉积物清晰、易于判断结果及免疫荧光淬灭时间长等优点。结论石蜡切片免疫荧光染色可作为冰冻切片免疫荧光染色的补充手段,它无需低温保存即能转运肾组织至肾脏病理诊断设备及水平较好的医院做免疫荧光检查,于国内推广具有现实意义。
Objective To explore the feasibility of immunofluorescent staining (IF) in formalin-fixed and paraffin-embedded kidney tissue. Methods The formalin- fixed and paraffin-embedded renal biopsy specimens of 10 patients with membranous nephropalhy, 25 IgA nephropathy and 10 lupus nephritis were selected for this study. First, the best working conditions including the deparaffinized duration with dimethylbenzene, the antigen restoring methods with enzyme or microwave, and the optima] concentrations of FITC-conjugated antibodies were investigated. Then, the direct IF was performed on the deparaffinized and antigen-restored sections (DARS) and compared to the direct IF on snap-frozen sections (SFS). Results In this study,the best deparaffinized duration with dimethylbenzene was 30 min, and the best antigen restoring method was incubation with pepsin (0.4%,pH2) for 10 min. The FITC-conjugated antibodies should be diluted to show the strongest specific fluorescence with weakest background of nonspecific fluorescence. The coincident rate of immunopathological diagnosis was 100% between the IF on the DARS and the SFS. In addition, the IF on DARS had the following benefits: easiness to judge results because of its better preserved tissue structure and clearer IF appearance; a longer IF fading time which will be propitious to observe IF results and take photographs. Conclusions The IF on DARS should become a complementary method of IF on SFS. Because the bother with long-distance transport of frozen tissue is avoided, it will be very convenient for the local hospitals to send the renal biopsy tissue to the big nephrology centers for IF examination, which has a greatly realistic significance in the current China.
出处
《中华肾脏病杂志》
CAS
CSCD
北大核心
2005年第6期315-319,共5页
Chinese Journal of Nephrology
