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MATERIALS AND METHODS Female ICR mice (Harlan, Houston, Tex.) weighing 18-23 g were ran- domly divided into 3 groups of 20 and given powdered feed (Purina Lab Chow 5002) supplemented with 0, 0.125, and 0.5% EQ HCI, respectively. Mice were killed by cervical dislocation, 4/group, after 2, 4, 6, and 10 wk of feeding; the liver, kidney, lung, and brain tissues were rapidly excised and weighed. Approximately 200 mg of each tissue sample with the ex- ception of the lung tissue was homogenized in 10 volumes (w/v) of ice- cold acetonitrile-water (7 : 3, v/v) with a Polytron (Brinckmann), then cen- trifuged for 10 min at 5000 x g. The clear supernatant was transferred into a screw-capped glass vial, centrifuged, then stored in a freezer (-20°C) for 2-3 h or until the two layers separated. Due to the small size of the lung tissue, most of it was homogenized in 20 volumes of solvent. The clear upper layers, approximately 60% of the total volume, contained most (>90%) of the EQ; therefore, only the upper layers were analyzed by HPLC detected by the fluoromonitor. In order to avoid possible diur- nal fluctuations, mice were killed at 8:00-10:00. Approximately 200 mg of another liver sample from each mouse was homogenized in 10 volumes (w/v) of ice-cold 0.25 M perchloric acid and centrifuged for 10 min at 5000 x g for the determination of GSH level. Three groups of mice, 4/group, were killed after 14 wk of feeding, 1/group/d from each feeding group, and the mitochondria were isolated from the liver, saving 2 samples of approximately 200 mg each to deter- mine the EQ residue and GSH levels. The liver mitochondria were iso- lated in 2.5 M sucrose buffer by differential centrifugaron Gohnson and Lardy, 1967) and the protein concentration was determined by the biuret method (Gornall et al., 1949). |
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8814402
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2楼2012-06-01 08:50:42
晴天1882
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爱与雨下: 金币+1 2012-06-03 21:04:38
zl0921516: 金币+15, 翻译EPI+1, ★★★★★最佳答案 2012-06-04 10:06:58
爱与雨下: 金币+1 2012-06-03 21:04:38
zl0921516: 金币+15, 翻译EPI+1, ★★★★★最佳答案 2012-06-04 10:06:58
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雌性ICR鼠(18-23g)随机分成三组,每组20只并正常喂养,饲料中分别添加0,0.125和0.5%的EQ HCI,在第2,4,6,10周的时候分别取每组中的四只老鼠断颈处死,迅速取出肝,肾,肺,脑组织并称重。大约200mg的组织(除去肺组织)悬浮于10倍体积预冷的乙腈-水(7:3体积比),然后5000g离心10min,上清液转移到有盖子的玻璃小瓶中,储存在-20°2-3h或者等到两相分离。由于肺组织比较小,一般悬浮于20倍体积的溶剂,清透的上清液大约占总体积的60%左右,包括EQ(超过90%),因此,只有上相才有高效液相分析,为了避免日常的波动,老鼠仔8点到十点之间处死。大约200mg肝组织悬浮于10倍体积预冷的0.25M过氯酸,5000g离心10min以达到检测谷胱甘肽水平。 第14周处死的老鼠,每组中取一只从肝组织分离线粒体,大约200mg来检测EQ和GSH水平,肝线粒体用2.5M蔗糖不同离心力来分离,用双缩脲法检测蛋白浓度。(太老的文献了) 大概是这样吧! |

3楼2012-06-03 18:01:46
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爱与雨下: , ? 2012-06-03 21:05:01
zl0921516: 金币+5, ★★★很有帮助 2012-06-04 10:07:08
| 雌性ICR小鼠(哈伦,休斯敦,得克萨斯州),体重18-23克然进行随机分为3组,每组20和粉状饲料(普瑞纳实验室周5002)辅以0,0.125,0.5%EQ人机交互,分别。颈椎脱位,4/group,后2,4,6,和10周处死小鼠,喂养;肝,肾,肺,脑组织迅速切除并称重。大约200毫克的每个组织样本前肺组织匀浆10卷(W / V)冰ception冷乙腈 - 水(7:3,V / V),然后与宝创(Brinckmann)中心trifuged为5000 XĞ10分钟。清澈的上清液转移到一个螺丝帽的玻璃瓶,离心,然后存放在冰箱(-20℃)为2-3小时,或直至两层分离。由于规模小肺组织,其中大部分被匀浆溶剂,在20卷。明确的上层,总量约60%,载大部分的EQ(> 90%),因此,只有上层进行了分析高效液相色谱法检测由fluoromonitor。为了避免可能diur信号波动,处死小鼠在8:00-10:00。约200毫克。另外从各组小鼠肝匀浆样品10卷冰冷的0.25 M过氯酸(W / V),离心10分钟G为5000 X GSH水平的决心。 三组小鼠,4/group,喂养14周后处死,1/group/d每次喂食组,线粒体分离从肝,节省约200毫克,每2个样品阻止挖掘情商残留和GSH水平。肝线粒体是ISO-lated差centrifugaron Gohnson和2.5 M蔗糖缓冲拉迪,1967年)和蛋白质浓度的缩二脲的测定方法(Gornall等,1949)。撤消修改Cíxìng ICR xiǎo shǔ (hā lún, xiūsīdūn, dekèsàsī zhōu), tǐzhòng 18-23 kè rán Jìnxíng suíjī fēn wéi 3 zǔ, měi zǔ 20 hé fěn zhuàng sìliào (pǔ ruì nà shíyàn shì Zhōu 5002) fǔ yǐ 0,0.125, 0.5% EQ rén jī jiāohù, fēnbié. Jǐngchuí tuōwèi, 4/group, hòu 2,4,6, hé 10 zhōu chǔsǐ xiǎo shǔ, Wèiyǎng; gān, shèn, fèi, nǎo zǔzhī xùnsù qiēchú Bìng chēng zhòng. Dàyuē 200 háokè de měi gè zǔzhī yàngběn qián Fèi zǔzhī yún jiāng 10 juǎn (W / V) bīng ception Lěng yǐjīng - shuǐ (7: 3, V/ V), ránhòu yǔ bǎo chuàng (Brinckmann) zhōngxīn Trifuged wèi 5000 X Ğ 10 fēnzhōng. Qīngchè de shàng qīng yè zhuǎnyí Dào yīgè luósī mào de bōlí píng, líxīn, ránhòu cúnfàng zài bīngxiāng ( -20℃) Wèi 2-3 xiǎoshí, huò zhízhì liǎng céng fēnlí. Yóuyú guīmó xiǎo Fèi zǔzhī, qízhōng dà bùfèn bèi yún jiāng róngjì, zài 20 juǎn. Míngquè de shàngcéng, zǒng liàng yuē 60% , zài Dà bùfèn de EQ (> 90% ), yīncǐ, zhǐyǒu shàngcéng jìnxíngle fēnxī Gāoxiào yè xiāng sèpǔ fǎ jiǎncè yóu fluoromonitor. Wèile bìmiǎn kěnéng diur Xìnhào bōdòng, chǔsǐ xiǎo shǔ zài 8:00-10:00. Yuē 200 háokè. Lìngwài cóng gè zǔ xiǎo shǔ gān yún jiāng yàngpǐn 10 juǎn Bīnglěng de 0.25 Mguò lǜ suān (W / V), líxīn 10 fēnzhōng G wèi 5000 X GSH shuǐpíng de juéxīn. Sān zǔ xiǎo shǔ, 4/group, wèiyǎng 14 zhōu hòu chǔsǐ, 1/Group/d měi cì wèishí zǔ, xiànlìtǐ fēnlí Cóng gān, jiéshěng yuē 200 háokè, měi 2 gè yàngpǐn zǔzhǐ Wājué qíngshāng cánliú hé GSH shuǐpíng. Gān xiànlìtǐ shì ISO- Lated chà centrifugaron Gohnson hé 2.5 M zhètáng huǎnchōng Lā dí,1967 nián) hé dànbáizhí nóngdù de suō èr niào de cèdìng Fāngfǎ (Gornall děng, 1949). |

4楼2012-06-03 19:00:16









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