摘要
用花生(Arachis hypogaea L.)种子的子叶组织作为模式材料,Epon 812包埋的组织进行厚切片后,按下列三步染色。第一步,PAS 反应:(1)在0.5%高碘酸(0.3%硝酸配制)中10分钟;(2)自来水洗1—2分钟,蒸馏水经过;(3)锡夫试剂中30分钟;(4)偏亚硫酸氢钠溶液三次洗涤,每次2分钟;(5)自来水洗5分钟,蒸馏水经过。第二步,苏丹黑 B 液染色:(1)70%酒精中1—2分钟;(2)新鲜配制的0.3%苏丹黑 B 液(70%酒精配制)中染色,40—60℃,30分钟至1小时;(3)70%酒精中分色1分钟;(4)自来水冼,蒸馏水经过。第三步,考马斯蓝染色:(1)7%醋酸中1—2分钟;(2)1%考马斯蓝(7%醋酸配制)中染色,60℃,20分钟;(3)0.1%醋酸液中分色1分钟;(4)自来水冼5分钟,蒸馏水经过;(5)自然干燥或在40℃电热温箱中烘干。干燥后的切片用甘油胶封固。经这三步染色的切片,细胞内的多糖、脂类和蛋白体同时被显示:淀粉粒及纤维素的细胞壁为樱红色,油滴为黑色,蛋白体为蓝色。制片能长期保存。
An effective cytochemical technique for the simultaneous demonstration of lipids,polysac- charities and protein bodies in the same section from the tissue embedded in Epon 812 is described.Thick sections of peanut cotyledon are used for a typical sample according to the following procelures.Firstly,PAS reaction:(1)Oxidize sections in 0.5% periodic acid in 0.3% nitric acid for 10 min,(2)Wash in running water for 1—2 min and then pass through distilled water,(3)Stain in Schiff's reagent for 30 min,(4)Wash in sodium metabisulfite 3 times,2 min for each time,(5)Wash in running water for 5 min and then pass through dis- tilled water.Secondly,Sudan black B staining:(1)Rinse section in 70% ethanol for 1—2 min, (2)Stain in fresh 1% Sudan black B in 70% ethanol for 30—60 min at 40—60℃,(3)Rinse in 70% ethanol for 1 min and then in distilled water.Thirdly,Coomassie brilliant blue R staining:(1)Rinse sections in 7% acetic acid for 1—2 main,(2)Stain in 1% Coomassie bril- liant blue R in 7% acetic acid for 20 min at 60℃,(3)Differentiate in 0.1% acetic acid for 1min,(4)Rinse in lunning water for 5 min and then pass through distilled water,(5)Dry at room temperature or in oven,40℃.The dry sections mount in glycerin-gelatin.After the above three step staining,the three main compounds of the cell can be stained simultane- ously.Starch grains and cellulose cell wall take cherry red colour,lipids appear in black, protein bodies are blue.The sealed slides can be kept permanently.
关键词
植物细胞化学
多糖
蛋白体
脂类
Cytochemistry
Polysaccharides
Protein bodies
Lipids
Thick resin section
