Toxoplasma gondii:: A paraformaldehyde-insensitive diaphorase activity acts as a specific histochemical marker for the single mitochondrion
Toxoplasma gondii:: A paraformaldehyde-insensitive diaphorase activity acts as a specific histochemical marker for the single mitochondrion
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DOI：
10.1006/expr.1998.4266
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发表时间：
1998-05-01
影响因子：
2.1
通讯作者：
Gross, U
中科院分区：
文献类型：
--
作者：
Seeber, F;Ferguson, DJP;Gross, U
Seeber，F.，Ferguson，DJP，and Gross，U. 1998.弓形虫电子显微镜，寄生虫的寄生虫，而宿主细胞刚地：一个多聚甲醛不敏感的心肌黄酶活性作为线粒体保持不染色，从而允许评价寄生虫特异性组织化学标记的单一寄生虫。实验-数字。Taling Parasitology 89，137-139. 1998学术出版社T.如前所述培养弓形虫速殖子。索引描述符和缩写：BSPT，2-（2-苯并噻唑基）-（Boothroyd等，1995 b）。使用的菌株为RH、PLK和RHβ1，5-苯乙烯基-3-（4-邻苯二甲酰肼基）四唑氯化物; DPI，二苯撑-其为能够表达Escherichia eiodonium chloride的RH的lacZ转染衍生物; DTNB，5-5-二硫代双-（2-硝基苯甲酸）;大肠杆菌β-半乳糖苷酶（Seeber和Boothroyd 1996）。使用EDTA，乙二胺四乙酸; EM，电子显微镜;感染接种在盖玻片上的人包皮成纤维细胞（HFF），24-HFF，人包皮成纤维细胞; ID，二苯基-HCl碘鎓; NBT，孔板，并在用4%多聚甲醛脱硝基蓝四唑固定之前生长18小时; NOS，一氧化氮合酶; PBS，在100 mM二甲胂酸钠缓冲液（pH 7.4）中的磷酸氢盐; PFA，多聚甲醛。温度在考察了无固定和固定20、40、60和120 min的效果后，得出了该最佳固定时间。在PBS中洗涤后，将样品在37 ℃下在含有50 mM Tris（pH 8）、1 mg/ml NADPH、0.2 mg/ml NBT和0.3%Triton X-100的溶液中孵育30分钟，所述溶液每次新鲜制备。然后将冲洗过的盖玻片固定在载玻片上，并用
Seeber, F., Ferguson, DJP, and Gross, U. 1998. Toxoplasma electron microscopy, the mitochondrion of the parasite while host cell gondii: A paraformaldehyde-insensitive diaphorase activity acts as a mitochondria remain unstained thus allowing evaluation of parasite specific histochemical marker for the single mitochondrion. Experimen- numbers. tal Parasitology 89, 137–139. 1998 Academic Press T. gondii tachyzoites were cultivated as described previously Index Descriptors and Abbreviations: BSPT, 2-(2-benzothiazolyl)-(Boothroyd et al. 1995b). The strains used were RH, PLK, and RHβ1, 5-styryl-3-(4-phtalhydrazidyl) tetrazolium chloride; DPI, diphenylen- which is a lacZ transfected derivative of RH able to express Escherichia eiodonium chloride; DTNB, 5-5-dithiobis-(2-nitrobenzoic acid); coli β-galactosidase (Seeber and Boothroyd 1996). Parasites were used EDTA, ethylanediaminetetraacetic acid; EM, electron microscope; to infect human foreskin fibroblasts (HFF) seeded on coverslips in 24-HFF, human foreskin fibroblasts; ID, iodoniumdiphenyl–HCl; NBT, well plates and grown for 18 h prior to fixation with 4% paraformaldenitroblue tetrazolium; NOS, nitric oxide synthase; PBS, phosphate- hyde in 100 mM sodium cacodylate buffer, pH 7.4, for 20 min at room buffered saline; PFA, paraformaldehyde. temperature. This optimum fixation time was arrived at after examining the effects of no fixation and 20, 40, 60, and 120 min fixation. After washing in PBS, samples were incubated for 30 min at 37C in a solution containing 50 mM Tris, pH 8, 1 mg/ml NADPH, 0.2 mg/ml NBT, and 0.3% Triton X-100 which was prepared fresh every time. The rinsed coverslips were then mounted on slides and viewed with