不同脱钙液对大鼠骨髓切片染色的影响

[目的]探讨不同脱钙液对Wistar大鼠胸骨骨髓切片染色效果的影响。[方法]取正常Wistar大鼠胸骨,采用10%中性福尔马林溶液充分固定,经流水冲洗后,分别浸入甲酸氯化铝脱钙液(A组)、5%甲酸脱钙液(B组)、5%硝酸脱钙液(C组)中进行充分脱钙后,再经流水冲洗,制作石蜡切片,HE染色,显微镜观察并评价其效果。[结果]①制片时间:5%甲酸组>甲酸氯化铝脱钙液>5%硝酸组;②HE染色效果:甲酸氯化铝脱钙液与5%甲酸组无显著差异,优于5%硝酸组。[结论]采用甲酸氯化铝脱钙液方法制作的大鼠胸骨骨髓切片,骨髓组织结构完整,细胞形态清晰,耗时较短,能够满足组织学评价的要求。     

适合北方地区冬季快速制备大鼠踝关节石蜡切片的脱钙方法

该研究旨在提供一种在较低室温下快速制作大鼠踝关节石蜡切片的方法,以期为大鼠关节炎的病理组织学评价提供重要技术支持。取类风湿关节炎模型(CIA)大鼠踝关节,经10%中性缓冲福尔马林溶液固定2 d,流水冲洗后,分为室温脱钙组和恒温脱钙组,两组均采用8%硝酸脱钙液脱钙,制作石蜡切片,采用HE染色法染色后,光镜下观察并评价其效果。结果表明,不同温度脱钙组制片后均能获得平整、染色良好的石蜡切片,可实现对关节的组织学评估,但恒温脱钙组脱钙完成时间明显短于室温脱钙组,可作为冬季较低室温条件下快速制备切片的脱钙方法。     

不同脱钙工艺对昆明小鼠骨组织石蜡切片效果的影响

为了获得最佳的昆明小鼠骨组织脱钙效果,以便进一步制得最佳的骨组织石蜡切片,采用四种脱钙工艺,即5%硝酸溶液脱钙22 h、8%硝酸溶液脱钙18 h、20%甲酸溶液脱钙26 h、Schmorls液脱钙16 h进行试验。结果表明:Schmorls液脱钙制得的切片H.E.染色效果最好,骨组织结构保留最完整;20%甲酸溶液脱钙的切片骨组织结构不完整,着色较弱;8%硝酸溶液脱钙的切片骨组织结构不清晰,对比度差;5%硝酸溶液脱钙的切片骨组织有皱褶,核浆对比不明显。说明用Schmorls液对昆明小鼠骨组织进行脱钙制得的切片质量最好。     

组织脱钙液对鹿角脱盘、鹿角柄的脱钙效果比较分析

为了明确鹿角脱盘和鹿角柄的较优脱钙方法,试验采用9种不同组织脱钙液对鹿角脱盘及鹿角柄进行脱钙,比较脱钙效果及组织形态和H.E.染色效果,综合分析筛选适合于鹿角脱盘、鹿角柄的脱钙方法。结果表明:在脱钙时间方面,采用脱钙液Ⅸ时间最短,脱钙液Ⅶ较短,采用脱钙液Ⅰ脱钙时间最长;在组织外观完整性方面,采用脱钙液Ⅰ脱钙后鹿角脱盘和鹿角柄外观最完整,脱钙液Ⅸ对组织损伤最大,脱钙液Ⅶ取相对完整的碎块组织制作了石蜡切片,切片视野清晰;在显微组织完整性方面,采用脱钙液Ⅰ和脱钙液Ⅶ脱钙后鹿角脱盘和鹿角柄组织较为完整;在H.E.染色效果方面,脱钙液Ⅰ的染色效果最好,脱钙液Ⅱ和脱钙液Ⅶ染色效果也较好。说明脱钙液Ⅶ对组织损伤小,脱钙效果好且脱钙时间短,切片染色效果较好,适合于常规条件下鹿角脱盘及鹿角柄脱钙;在脱钙时间充裕的情况下建议选择脱钙液Ⅰ对鹿角脱盘和鹿角柄进行脱钙,对组织损伤最小,脱钙效果最好,切片染色效果也好。     

Masson三色染色与HE染色在豚鼠牙齿组织石蜡切片中的比较研究及应用

针对牙齿组织形态学研究的难点,选择适宜固定液,用EDTA-2Na进行脱钙,完成了牙齿组织石蜡切片的制作。通过确定染色程序及各种染料的最佳作用时间,分别利用Masson三色染色法、传统的苏木精和伊红（HE）染色法对牙齿组织切片进行了染色观察。染色结果表明,与HE染色法相比,Masson三色染色更能使牙齿的各主要成分以不同颜色显现出来,色彩对比鲜明,亮丽润眼,为牙齿组织生理及病理组织形态学研究奠定了基础。针对机体中硬组织的相似特性,该试验方法可以在骨骼、软骨等硬组织的形态学研究中参考、改进并推广使用。     

快速HE染色切片制作

苏木精和伊红染色方法 (简称HE染色 )是生物学和医学领域中组织学和细胞学等学科中最常见的切片染色方法。染色质量的好坏直接影响到细胞观察的科学性和病理学诊断的准确性。但在一些特殊情况下 ,在保证切片质量的前提下需要进行快速石蜡HE染色切片。经过长时间的石蜡切片HE染色的制作实践 ,对实验室不具备自动脱水机和自动染色机的HE染色切片快速制作方法进行了如下的总结。1　石蜡切片的快速制作常规 (普通 )快速包埋法 :此方法快速而且制片的效果很好 ,整个过程一般在 10～ 15min完成。过程如下 :①取材、固定、脱水 :取材小而薄 ,组…     

新西兰白兔毛囊体外培养条件优化及其增殖能力的研究

为建立新西兰白兔毛囊体外培养模型,探究其最适的培养环境,本实验选用2月龄新西兰白兔,采集触须毛囊,分离组织和脂肪,保留完整毛囊。将完整无损的毛囊放入24孔板,每孔1根,加入William’s E、DMEM、MEM3种不同培养基,每组6个重复,每2d在倒置显微镜下观察毛囊毛干的生长,并利用HE和PCNA免疫荧光染色检测毛囊的形态和增殖能力。结果显示：3种培养基中毛囊毛干在体外培养4d后生长减缓,在2 d与4 d时的生长速度均大于6 d(P<0.05)。其中,William’s E组生长速度高于MEM组（P<0.05）,且William’s E组生长最快。4 d时,HE染色发现William’s E组毛囊结构完整,毛乳头清晰,而此时DMEM和MEM组毛乳头已经消失。进一步用PCNA免疫荧光染色,发现Williams’E组PCNA在毛乳头阳性表达,DMEM和MEM组毛囊表达仅表达在外根鞘。因此,William’sE培养基最适宜新西兰白兔触须毛囊生长,毛乳头增殖能力最强。     

三个品种肉兔人工感染兔巴氏杆菌的抗病性比较研究

分别用2个和4个最小致死剂量的巴氏杆菌强毒人工感染福建黄兔、新西兰兔和比利时兔,以比较这三个肉兔品种对巴氏杆菌急性感染是否存在一定抗病性差异。结果:福建黄兔、比利时兔和新西兰白兔高、低剂量组的发病率均为100%,除新西兰白兔低剂量组死亡率略低(90%)外,其余各组死亡率均为100%。从各组死亡时间来看,低剂量组的福建黄兔、比利时兔和新西兰白兔攻毒后2天内死亡率分别为80%、100%和90%;高剂量组的比利时兔24h内全部死亡,而福建黄兔和新西兰白兔24h内死亡率分别为80%(8/10)和70%(7/10)。本试验结果表明:福建黄兔、新西兰白兔和比利时兔在对抗巴氏杆菌强毒人工感染急性发病时,其抗病力没有差异。     

两种固定液对鸭小肠肥大细胞染色效果的比较

为了探讨不同固定液对鸭小肠肥大细胞甲苯胺蓝染色效果的影响及鸭小肠内肥大细胞的数量和分布情况，试验选择5只100日龄的健康淮南麻鸭公鸭为研究对象，取鸭只十二指肠、空肠和回肠组织样品，均分为两份，分别置于10%中性福尔马林固定液和卡恩氏(Carnoy)固定液中固定，常规制作石蜡切片，切片经甲苯胺蓝染色后，观察小肠肥大细胞染色效果和肥大细胞形态、数量及分布，并进行统计分析。结果表明：Carnoy固定液对肥大细胞的染色效果优于10%中性福尔马林固定液，染色评分极显著高于10%中性福尔马林固定液(P<0.01);Carnoy固定液固定的十二指肠、空肠、回肠肥大细胞数量均极显著高于10%中性福尔马林固定液(P<0.01)。鸭十二指肠、空肠和回肠中肥大细胞形态和数目不同，在黏膜固有层血管、腺管周围的结缔组织中分布较多；空肠中肥大细胞数量最多，形态多样；十二指肠和回肠中肥大细胞数量递减，形态多为圆形。说明与10%中性福尔马林固定液相比，经Carnoy固定液固定的鸭小肠组织染色后，可清晰显示肥大细胞的形态分布。     

海豹主要器官的组织结构特点

1.1 材料 雄性海豹3只(饲养于长春市胜利公园和广州动物园),死后迅速取其内脏器官,切成小块,分别固定于10%福尔马林、无水酒精和Bouin氏液中.1.2 方法 材料经固定后,制成石蜡切片,HE染色.另外,还对胃肠采用PAS染色,对胃肠、肾上腺采用Masson氏银浸法染色,对胰腺采用Gomori醛复红法染色.切片在光镜下与普通哺乳动物相应器官进行对比观察,记录其组织结构特点.     

日粮中缺硒对鸡空肠肥大细胞组织化学和超微结构的影响

为探讨硒缺乏对鸡空肠中肥大细胞(MC)的形态学和超微结构的影响,将10只鸡随机分成低硒试验组(饲喂低硒日粮0.032 mg/kg)和正常对照组,在75日龄时断头处死立即取出空肠,进行甲苯胺蓝、长时间甲苯胺蓝、阿利新蓝-藏花红染色、阿利新蓝-高碘雪夫氏反应和醛品红-阿利新蓝5种染色并进行透射电镜观察.结果显示,采用不同染色方法所获得的MC数量、大小、形态、颜色及颗粒的黏多糖成分均不相同.电镜下,MC细胞胞浆内充盈大量的特征性颗粒,颗粒内容物呈均质状,电子密度高低不等,核圆形或椭圆形,胞质中可见高尔基复合体和较多的线粒体;试验组可见MC数量显著增加并与变性细胞紧密相邻,异染色质增多,核偏位或固缩,胞质稀疏,同时可见MC内颗粒内容物释放后的空隙或空泡.因此,组化染色提示肥大细胞存在不同亚型;试验组MC数量明显增加,这提示MC是重要的免疫细胞之一,在机体的免疫调节中发挥着重要的作用.     

Acid mucopolysaccharides in mammary tumors of dogs.

The predominant acid mucopolysaccharides found in selected epithelial mammary tumors of dogs stained with alcian blue and were labile to hyaluronidase digestion. These histochemical characteristics identified them as hyaluronic acid, chondroitin-4- and chondroitin-6-sulfate. The intensity of the staining of these acid mucopolysaccharides varied in a transitionary process from a precartilaginous to a pseudocartilaginous intercellular matrix to mature hyaline cartilage. The tumor acid mucopolysaccharides were indistinguishable from those associated with formation of cartilage in developing mammals; such cartilage is reported to be produced only by cells of mesodermal origin. There was no evidence to suggest transitional changes in myoepithelial cells, neoplastic epithelial cells or their components that could contribute to the formation of the acid mucopolysaccharides. It was concluded that the heterotopic tissues (cartilage, bone and fibrous connective tissue) in the epithelial mammary tumors were derived from cells of mesodermal origin and formed the adjacent stroma in areas of neoplasia.     

透明质酸钠与软骨下钻孔对兔软骨损伤修复的对比试验

试验通过建立兔部分软骨损伤模型,采用外源性注射透明质酸钠和软骨下钻孔两种治疗方法,比较两种方法的修复效果及早期修复机制,为临床应用提供试验依据。试验选用36只哈白兔建立膝关节软骨损伤模型后随机均分为3组：自愈组（阴性对照）、用药组（透明质酸钠注射）、钻孔组（软骨下钻孔）。术后1、4、6、8周,采用RT-PCR方法检测修复物中Ⅱ型胶原（Col Ⅱ）、蛋白聚糖（Agg）、Ⅹ型胶原（Col Ⅹ）、Ⅰ型胶原（Col Ⅰ）及Runx2的mRNA表达量;术后8周,制作软骨切片进行HE染色、番红-固绿染色和甲苯胺蓝染色,分析3组的修复效果及修复机制。结果显示,两试验组软骨损伤都被明显修复,染色结果显示修复组织中软骨细胞增多,具有软骨典型的陷窝样结构,用药组阳性着色优于钻孔组。用药组软骨的特异性基因Agg和Col Ⅱ的mRNA表达量显著高于钻孔组（P < 0.05）,软骨细胞肥大相关基因Col Ⅹ和Runx2的表达量显著低于钻孔组（P < 0.05）。自愈组Col Ⅰ的mRNA表达量最高,钻孔组次之,用药组最低。试验结果表明,用药及钻孔对关节软骨损伤都有不同程度的治疗修复作用,自愈组修复物成分主要为纤维组织,透明质酸钠治疗后产生了较多的透明软骨成分,而钻孔修复后的表层覆盖薄层纤维软骨成分,在骨缺损短期修复中,透明质酸钠的修复效果优于钻孔疗法。     

小鼠软骨发育的形态学观察

应用茜素红—阿利新蓝双重染色技术制作骨与软骨透明标本,观察幼龄小鼠软骨发育的形态学变化。取1、3、5、7、9、15日龄完整幼鼠,水淹致死后去除皮肤、眼、脑、内脏和脂肪组织,乙醇固定,丙酮脱脂,自来水冲洗,茜素红—阿利新蓝混合染色,氢氧化钾透明,各梯度甘油脱水,纯甘油保存。结果小鼠骨骼双重染色硬骨显示为红色,软骨显示为蓝色。随着小鼠日龄的增长,小鼠肋软骨的骨化最为明显,后肢长骨末端的骺软骨板和髋部软骨骨化过程清晰可见。此试验可以清晰的显示不同日龄小鼠肋软骨、后肢关节软骨、髋部软骨的骨化过程。     

Evaluation of intra-articularly administered sodium monoiodoacetate-induced chemical injury to articular cartilage of horses.

Three doses of sodium monoiodoacetate (MIA) were used to induce degenerative changes in articular cartilage in middle carpal joints of horses. Twelve young (2- to 5-year-old) horses, free of lameness, were randomly allotted to 3 groups. One middle carpal joint of each horse was injected with 0.9% NaCl solution (control joint). The contralateral middle carpal joint was injected with 0.09 mg of MIA/kg of body weight (group 1); 0.12 mg/kg (group 2); or 0.16 mg/kg (group 3). After MIA administration, horses were allowed ad libitum exercise in a 2-acre paddock for 12 weeks. At the end of the study, gross and microscopic tissue changes were evaluated and biochemical analyses of articular cartilage were done. Grossly, diffuse partial-thickness articular cartilage lesions were observed in group-2 (n = 2) and group-3 (n = 4) horses, but not in group-1 horses. Articular cartilage uronic acid content was significantly (P less than 0.03) decreased in all MIA-injected joints, compared with controls. Articular cartilage matrix staining with safranin-O was decreased in 3 of 4 MIA-injected joints of group-1 horses and in all MIA-injected joints of group-2 and group-3 horses, compared with controls (P less than 0.06). Microscopic degenerative changes in articular cartilage were not significantly different between MIA-injected and control joints in group-1 horses, but were increased (P less than 0.06) in all MIA-injected joints of group-2 and group-3 horses, compared with controls. Qualitatively, decreased matrix staining and degenerative changes were more severe in group-3 horses. On the basis of articular cartilage gross and microscopic changes, as well as biochemical changes, 0.12 mg of MIA/kg injected intra-articularly was determined to induce moderate degrees of articular cartilage degeneration. This model of chemically induced articular cartilage injury could be useful for evaluating treatment effects of anti-arthritic drugs in horses.     

日粮中缺硒对鸡空肠肥大细胞组织化学和超微结构的影响

为探讨硒缺乏对鸡空肠中肥大细胞(MC)的形态学和超微结构的影响,将10只鸡随机分成低硒试验组(饲喂低硒日粮0.032 mg/kg)和正常对照组,在75日龄时断头处死立即取出空肠,进行甲苯胺蓝、长时间甲苯胺蓝、阿利新蓝-藏花红染色、阿利新蓝-高碘雪夫氏反应和醛品红-阿利新蓝5种染色并进行透射电镜观察。结果显示,采用不同染色方法所获得的MC数量、大小、形态、颜色及颗粒的黏多糖成分均不相同。电镜下,MC细胞胞浆内充盈大量的特征性颗粒,颗粒内容物呈均质状,电子密度高低不等,核圆形或椭圆形,胞质中可见高尔基复合体和较多的线粒体;试验组可见MC数量显著增加并与变性细胞紧密相邻,异染色质增多,核偏位或固缩,胞质稀疏,同时可见MC内颗粒内容物释放后的空隙或空泡。因此,组化染色提示肥大细胞存在不同亚型;试验组MC数量明显增加,这提示MC是重要的免疫细胞之一,在机体的免疫调节中发挥着重要的作用。     

Histochemical study of normal and collapsed tracheas in dogs

Tracheas from 15 toy breed dogs with normal tracheas and 6 dogs with collapsed tracheas were examined histologically and histochemically. Tracheal cartilage was analyzed for chondroitin sulfate by means of alcian blue (CEC method) and for calcium with alizarin red S. Cartilage arcs from dogs with collapsed tracheas had areas that were apparently hypocellular, and some had other areas that appeared like fibrocartilage or fibrous tissue. Histochemically, collapsed tracheal cartilage had less chondroitin sulfate and calcium than did normal tracheal cartilage. Cartilage arcs from the collapsed tracheas were not uniformly affected to the same degree, and parts of a given tracheal arc appeared normal, whereas other parts had an abnormal histologic appearance.     

Quantitative microanalysis of equine synovial fluid glycosaminoglycan concentration

An alcian blue precipitation method for quantifying the hyaluronic acid (HA) and sulphated glycosaminoglycan concentration (SGAG) in solutions containing both compounds was assessed. The assay was found to be rapid and reliable in solutions containing 0 to 200 mg of HA/dl and 50 to 1,000 micrograms of SGAG/dl, and was not affected by the presence of protein, hemoglobin, or methemoglobin in concentrations normally found in synovial fluid. The HA and SGAG concentrations in intercarpal synovial fluid from 13 clinically normal and 11 arthritic horses were evaluated. A relationship was not found between the concentration of HA and SGAG and any other synovial fluid variable. The SGAG concentration was found to be markedly high in several of the synovial fluid samples from arthritic horses, but did not correlate with the degree of articular cartilage erosion.     

The Effect of Trans-Stifle External Skeletal Fixation and Hyaluronic Acid Therapy on Articular Cartilage in the Dog

Transarticular external skeletal (TES) fixators were applied unilaterally to the stifle joints of 10 young adult dogs. After 4 weeks, the fixators were removed from all dogs. Two dogs were not allowed a remobilization period, whereas 8 dogs were provided with 4 additional weeks of weight-bearing activity in a kennel run. Four dogs were given high-molecular weight hyaluronic acid by intra-articular injection weekly during the remobilization period. Clinical gait evaluations and range of motion were determined during the remobilization period. Articular cartilage samples from both stifle joints of all dogs were evaluated histologically and histochemically. No significant differences in gait scores or range of motion were noted between treated and untreated dogs. Articular cartilage proteoglycan content was reduced after 4 weeks of trans-stifle external skeletal fixation as determined by loss of alcian blue (AB) histochemical staining. Improved homogeneity of histochemical staining was observed after remobilization. However, remobilization was associated with histological damage to the surface and tangential layers of articular cartilage. Remobilization combined with hyaluronic acid (HA) therapy improved histochemical staining and reduced structural damage to articular cartilage when compared with remobilization alone.     

Mast cells of the bovine trachea: staining characteristics, dispersion techniques and response to secretagogues.

Sections of the lower trachea of cattle, fixed in either Carnoy''s or formalin, were stained with toluidine blue, alcian blue, or alcian blue and safranin O to study the mast cell population. After toluidine blue staining, about twice as many cells in tissue fixed in Carnoy''s contained dark blue granules compared with tissue fixed in formalin. In addition, for the first time in cattle, a population of cells containing red granules was identified after staining with alcian blue and safranin O. Most of these red granules were formalin sensitive. An enzymatic dispersal technique for mast cells is described that yielded 9.4+/-0.4% mast cells (percentage of nucleated cells) with a viability of 92.3+/-0.6%. Spontaneous histamine release was 3.3+/-0.8%. Dispersed mast cells were challenged with various immunological and nonimmunological secretagogues. The calcium ionophores, A23187, ionomyocin, and BrX537A, were effective in releasing up to 94% of histamine in mast cells in a dose-response relationship. Pasteurella haemolytica culture supernate caused about 10% histamine release at a dose of 0.5 mg/mL after correction for spontaneous release. The average histamine content of the mast cells was 6.6+/-1.0 pg/cell. Cytospins of dispersed cells fixed in Carnoy''s and stained with alcian blue and safranin O contained mast cells with blue and red granules, and a few cells with a mixture of both granule types. Based on the effects of type of fixation, staining characteristics and histamine content, a mix of subtypes of mast cells is present in the bovine trachea. However, functionally they respond to secretagogues differently than rodent mast cells. Without an immunological secretagogue, studies to determine compounds that will be effective in blocking mast cell degranulation will be limited.