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引自链接:http://www.flickr.com/photos/28462956@N06/3341839938/in/photostream/
100%(w/v)三氯乙酸的配制方法:
500g三氯乙酸用227ml水来溶解,所得溶液即100%三氯乙酸溶液。避光,4度保存。
用TCA去除蛋白:
1) To a sample of protein solution add Trichloroacetic acid (TCA) 100% to get 13% final concentration. Mix and keep 5min at –20 ℃ and then 15min 4℃; or longer time at 4℃ without the –20℃ step for lower protein concentration.
(preparation of 100% TCA: 454ml H2O/kg TCA. Maintain in dark bottleat 4oC.Be careful, use gloves!!!).
用三氯乙酸沉淀蛋白的工作浓度为TCA终浓度13%
2) Spin 15min 4oC in microfuge at maximum speed (15000g).
或者:(方法二)
1. TCA precipitation
Before loading on SDS-PAGE, proteins can be precipitated by mixing 20% TCA with protein samples (1:1, 10% TCA final concentration). Keep samples in ice 20 min. Then centrifuge the samples 10 min at 16.000 x g. The protein pellet is then washed with ethanol/ethyl acetate (1:1) and centrifuged 10 min; 16 000 x g, three times. The final protein pellets is speed-vac dried dissolved in 2X Laemmli buffer at 100 °C for 15 min.
2. Acetone precipitation
Protein samples are mixed with acetone for a final acetone concentration of 80%. Keep samples in ice 20 min. Then centrifuge the samples 10 min at 16.000 x g. The protein pellets is speed-vac dried dissolved in 2X Laemmli buffer at 100 °C for 15 min.
相关链接:http://www.hos.ufl.edu/meteng/HansonWebpagecontents/Proteinprotocols.html
引自链接:http://blog.bioon.com/uploadfiles/2008-4/7232372059.jpg
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