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Polychrome staining of epoxy semithin sections using cacodylic buffer as a stain solvent. 用羧酸缓冲液作染色溶剂对环氧树脂半薄切片进行多色染色。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106989
S Pasyk, W Bartosik, A Fabry
Many polychromatic stains are in use for epoxy-embedded tissues (Horobin 1983). We should like to report a quick and easy polychromatic staining procedure that we find useful for routine use. Formerly the stain we used was prepared in 20 ml water and 5 ml 96% alcohol, and gave polychromatic staining only at pH 7.4 obtained by the addition of 1 N NaOH. However, the stain gave satisfactory results only for two or three days. We found that stabilization of the staining solution through the use of an ethyl alcohol-cacodylic buffer solvent increased the life of the staining solution. This was convenient because the cacodylic buffer is used in our laboratory as a component of fixatives, and is not prepared specially for the staining.
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引用次数: 1
A 595 nanometer band-pass filter enhances the contrast of in situ hybridization signals on chromosome observed after biotin/avidin-alkaline phosphatase localization. 595纳米带通滤波器增强了生物素/亲和素碱性磷酸酶定位后染色体原位杂交信号的对比度。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106986
H Yasue, T Awata, S Muramatsu

In situ hybridization with DNA probes labeled with biotin and detected by the avidin-alkaline phosphatase/5-bromo-chloroindoxyl phosphate-nitro blue tetrazolium system has been used to localize DNA sequences in chromosomes. To observe the hybridization signals, a phase contrast microscope has often been used because of the good visibility it provides. Use of a 595 nm band pass filter with the phase contrast microscope enhances signal contrast after in situ hybridization without reducing resolution.

用生物素标记的DNA探针和亲和素-碱性磷酸酶/5-溴-氯吲哚磷酸-硝基蓝四氮唑体系检测的原位杂交技术已被用于定位染色体中的DNA序列。为了观察杂交信号,经常使用相衬显微镜,因为它提供了良好的可视性。使用595 nm带通滤波器与相衬显微镜增强原位杂交后的信号对比度而不降低分辨率。
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引用次数: 1
An improved flat embedding technique for immunoelectron microscopy. 一种改进的免疫电子显微镜平面包埋技术。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106987
S M Yu, I R Schwartz

Immunocytochemical staining has been widely used for localizing various hormonal antigens, protein markers and putative neurotransmitters in tissues. Immunostained sections can be examined light microscopically and specific areas selected for electron microscopic study.

免疫细胞化学染色已广泛应用于组织中各种激素抗原、蛋白质标记物和假定的神经递质的定位。免疫染色切片可以在光镜下检查,并选择特定区域进行电镜研究。
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引用次数: 11
Note from the Biological Stain Commission: certification of Wright stain solution. 来自生物染色委员会的说明:莱特染色液的认证。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106991
E A Schenk, C T Willis
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引用次数: 5
An improved method of making tissue carriers for TEM and SEM fixation. TEM和SEM固定用组织载体的改进制备方法。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106990
J F Pilger
Hazards in fixing small pieces of tissue for electron microscopy include damage, drying, or loss. Over the years, microstrainer tissue carriers have been developed to minimize these problems. Construction materials have included glass tubing, copper grids for electron microscopy, stainless steel screen, and bolting silk (Padawer 1951, Friend 1963, Bronskill 1970). Carriers made from plastic embedding molds (e.g., BEEM capsules) with either TEM grids attached to the conical tip (Buchanan 1965) or Nitex screen cloth held to one end by a retaining ring have proven to be inexpensive and popular, though the former has a very small filtration area and in the latter small tissues may be lost or crushed between the screen cloth and the bottom rim of the carrier. This note describes a carrier in which Nitex is permanently sealed to the bottom edee of a BEEM capsule cylinder.
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引用次数: 0
A method for preparing and staining histological sections containing titanium implants for light microscopy. 一种制备和染色光学显微镜用含钛植入物的组织学切片的方法。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106984
K Gotfredsen, E Budtz-Jörgensen, L N Jensen

An improved method for preparing and staining ground tissue-implant sections for light microscopy is presented. Undecalcified tissue blocks with titanium implants were dehydrated in an ascending series of ethanol and stained in toto with basic fuchsin. Specimens were infiltrated and embedded in methyl methacrylate and sections were prepared using a cutting-grinding-system. The polished surface was counterstained with light green or anilin blue. Light polymerizing resin was used as slide mounting medium and for mounting the coverglass. The sections obtained were 10-15 microns thick with tissue architecture which clearly differentiated structures at the tissue-implant interface. The method was very useful for computer assisted morphometric analysis.

提出了一种光镜下制备和染色地面组织植入切片的改进方法。带钛植入物的未钙化组织块在递增系列乙醇中脱水,并用碱性品红染色。将标本浸润并包埋在甲基丙烯酸甲酯中,并用切割-研磨系统制备切片。抛光后的表面涂上了淡绿色或动物蓝。采用轻聚合树脂作为载玻片的贴装介质和盖板玻璃的贴装。获得的切片厚度为10-15微米,组织结构清晰,组织-植入物界面处结构分化明显。该方法对计算机辅助形态计量学分析非常有用。
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引用次数: 79
Lymphocyte membrane antigens in glycol methacrylate embedded tissue. 甲基丙烯酸乙二醇包埋组织中的淋巴细胞膜抗原。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106982
V Glezerov, B Dorsett

The use of formalin or Michel's solution either alone or in combination with acetone, and acetone, methanol or ethanol alone as fixatives, and glycol methacrylate as embedding medium were evaluated for their suitability in procedures to detect lymphocyte membrane antigens by OKT and Leu monoclonal antibodies in human tonsils. No staining was detected in sections fixed in 70% or absolute ethanol and embedded in glycol methacrylate with either the direct immunofluorescence or avidin-biotin methods. Fixation in Michel's solutions plus acetone at room temperature revealed staining by both. Neither method resulted in staining after fixation in Michel's solution plus acetone at 4 C presumably due to the slow action of the fixative. Staining was enhanced using a combination of primary and secondary biotinylated antibodies. Dual staining allowed concurrent detection of two antigens in the same section. Glycol methacrylate embedding is a possible replacement for ultracold storage in the preservation of tissue for immunofluorescent staining.

使用福尔马林或米歇尔溶液单独或与丙酮联合,丙酮、甲醇或乙醇单独作为固定剂,甲基丙烯酸乙二醇酯作为包埋介质,评估其在通过OKT和Leu单克隆抗体检测人扁桃体淋巴细胞膜抗原的程序中的适用性。采用直接免疫荧光法或亲和素-生物素法对70%或无水乙醇固定、甲基丙烯酸乙二醇酯包埋的切片均未见染色。在米歇尔溶液和丙酮中室温固定,显示两者都染色。这两种方法在米歇尔溶液加丙酮中固定后都没有染色,可能是由于固定液的作用缓慢。使用一级和二级生物素化抗体的组合增强了染色。双重染色允许在同一切片中同时检测两种抗原。甲基丙烯酸乙二醇包埋是一种可能的替代超冷储存保存组织用于免疫荧光染色。
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引用次数: 1
A versatile new mineralized bone stain for simultaneous assessment of tetracycline and osteoid seams. 一种多功能的新矿化骨染色剂,用于同时评估四环素和类骨接缝。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106985
A R Villanueva, K D Lundin

A versatile mineralized bone stain (MIBS) for demonstrating osteoid seams and tetracycline fluorescence simultaneously in thin or thick undecalcified sections has been developed. Bone specimens are fixed in 70% ethanol, but 10% buffered formalin is permissible. Depending upon one's preference, these specimens can be left unstained or be prestained before plastic embedding. Osteoid seams are stained green to jade green, or light to dark purple. Mineralized bone matrix is unstained or green. Osteoblast and osteoclast nuclei are light to dark purple, cytoplasm varies from slightly gray to pink. The identification of osteoid seams by this method agrees closely with identification by in vivo tetracycline uptake using the same section from the same biopsy. The method demonstrates halo volumes, an abnormal, lacunar, low density bone around viable osteocytes in purple. This phenomenon is commonly seen in vitamin D-resistant rickets, fluorosis, renal osteodystrophy, hyperparathyroidism, and is sometimes seen in fluoride treated osteoporotic patients. In osteomalacic bone, most osteoid seams are irregularly stained as indicated by the presence of unmineralized osteoid between mineralized lamellae. The method has been used effectively in staining new bone formation in hydroxyapatite implants and bone grafts. Old, unstained, plastic embedded undecalcified sections are stained as well as fresh sections after removal of the coverslip. This stain also promises to be valuable in the study of different metabolic bone diseases from the point of view of remodeling, histomorphometry, and pathology.

开发了一种多功能矿化骨染色(MIBS),用于同时在薄或厚未钙化切片上显示类骨接缝和四环素荧光。骨标本固定在70%的乙醇中,但10%的缓冲福尔马林是允许的。根据个人喜好,这些标本可以不染色,也可以在塑料包埋前进行染色。骨样接缝呈绿色至翡翠绿色,或浅至深紫色。矿化后的骨基质无色或呈绿色。成骨细胞和破骨细胞细胞核为浅紫色至深紫色,细胞质为浅灰色至粉红色。用这种方法鉴定类骨缝与用同一活检组织的同一切片进行体内四环素摄取鉴定非常一致。该方法显示晕状体积,紫色为活骨细胞周围的异常腔隙低密度骨。这种现象常见于维生素d抗性佝偻病、氟中毒、肾性骨营养不良、甲状旁腺功能亢进,有时也见于氟化物治疗的骨质疏松症患者。在骨软化性骨中,由于矿化片之间存在未矿化的类骨,大多数类骨接缝呈不规则染色。该方法已在羟基磷灰石种植体和骨移植体的新骨形成染色中得到了有效的应用。旧的,未染色的,塑料嵌入的未钙化的部分被染色,以及在去除盖盖后的新鲜部分。该染色也有望从重塑、组织形态学和病理学的角度研究不同的代谢性骨疾病。
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引用次数: 98
Thiéry test in combination with a uranyl acetate-lead citrate staining. 硫胺嘧啶试验结合醋酸铀酰-柠檬酸铅染色。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106988
M Weber
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引用次数: 5
Effect of cytochrome c concentration on the ultrastructural appearance of bovine nasal cartilage proteoglycans. 细胞色素c浓度对牛鼻软骨蛋白聚糖超微结构外观的影响。
Pub Date : 1989-05-01 DOI: 10.3109/10520298909106983
P Front, C Dauguet, D R Mitrovic

Bovine nasal cartilage proteoglycan monomers were studied by Kleinschmidt and Zahn's molecular spreading technique as modified by Rosenberg et al. By decreasing the cytochrome c concentration in the epiphase to 2 micrograms per 100 microliters we were able on nitrocellulose-coated grids routinely to obtain highly contrasted and well spread proteoglycan monomers with a characteristic brush-like appearance and, sometimes, a clearly distinguishable hyaluronic acid binding region. Previously, a hyaluronic acid binding region has only been observed routinely in spread proteoglycan aggregates, and a brush-like structure of proteoglycan monomers on carbon-coated grids, but with considerably less precision due to the poor contrast of the molecules. Molecular spreading was further improved by decreasing the cytochrome c concentration in the epiphase to less than 2 micrograms per 100 microliters, but contrast was reduced making visualization of molecular details difficult.

采用Kleinschmidt和Zahn的分子扩散技术对牛鼻软骨蛋白多糖单体进行了研究,并经Rosenberg等人改良。通过将细胞色素c浓度降低到每100微升2微克,我们能够在硝化纤维素包覆的网格上常规地获得高度对比和均匀分布的蛋白多糖单体,具有典型的刷子状外观,有时,透明质酸结合区域清晰可辨。在此之前,透明质酸结合区域仅在分散的蛋白聚糖聚集体和碳涂层网格上的蛋白聚糖单体的刷子状结构中常规观察到,但由于分子对比度差,其精度相当低。通过将细胞色素c浓度降低到每100微升2微克以下,进一步改善了分子扩散,但对比度降低,使分子细节的可视化变得困难。
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引用次数: 3
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Stain technology
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