Improved technique utilizing nonfat dry milk for analysis of proteins and nucleic acids transferred to nitrocellulose.
Improved technique utilizing nonfat dry milk for analysis of proteins and nucleic acids transferred to nitrocellulose.
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DOI：
10.1016/0735-0651(84)90049-9
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发表时间：
1984-01-01
期刊：
影响因子：
--
通讯作者：
Elder, J. H.
中科院分区：
文献类型：
--
作者：
Johnson, D. A.;Gautsch, J. W.;Elder, J. H.
描述了一种简单、经济和有效的方法，用于分析转移到硝酸纤维素上与抗体或核酸探针反应的蛋白质（Western印迹法）和DNA（Southern印迹法）。该技术利用脱脂奶粉作为蛋白质-核酸来源，用于阻断非特异性反应，作为孵育介质，并用于随后的洗涤以除去未反应的试剂。孵育混合物，称为BLOTTO（牛乳糖转移技术优化剂），是上级牛血清白蛋白（BSA）或明胶，以防止非特异性吸收的蛋白质印迹分析，并不需要洗涤剂或离液剂，以有效地降低背景。BLOTTO在氯化钠-柠檬酸钠中适当稀释后，在Southern印迹分析中与后一种技术中通常使用的更复杂的混合物一样有效。相对于正常BSA载体，BLOTTO也很好地用于封闭、孵育和洗涤ELISA板测定，为实验室节省了相当多的费用。
A simple, economical and efficient procedure for analysis of proteins (Western blotting) and DNA (Southern blotting) transferred to nitrocellulose for reaction with antibodies or nucleic acid probes is described. The techniques utilize dried skim milk as a protein-nucleic acid source for blocking non-specific reactions, as an incubation medium, and for subsequent washing to remove unreacted reagents. The incubation cocktail, termed BLOTTO (Bovine Lacto Transfer Technique Optimizer), was superior to bovine serum albumin (BSA) or gelatin for preventing non-specific absorption in Western blot analyses and did not require detergents or chaotropic agents to effect efficient decrease of background. BLOTTO, at the proper dilution in NaCl-Na citrate, was as efficient in Southern blot analyses as more complicated cocktails typically used in the latter technique. BLOTTO also worked well for blocking, incubating and washing ELISA plate assays relative to the normal BSA carrier, at considerable savings to the laboratory.