{"id":407,"date":"2016-05-30T00:00:00","date_gmt":"2016-05-30T00:00:00","guid":{"rendered":"https:\/\/www.thermofisher.com\/blog\/learning-at-the-bench\/optimization-of-affinity-purification\/"},"modified":"2025-03-10T10:29:23","modified_gmt":"2025-03-10T01:29:23","slug":"optimization-of-affinity-purification","status":"publish","type":"post","link":"https:\/\/www.thermofisher.com\/blog\/learning-at-the-bench\/optimization-of-affinity-purification\/","title":{"rendered":"\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u7cbe\u88fd\u306b\u304a\u3051\u308b\u7d50\u5408\u30fb\u6eb6\u51fa\u6761\u4ef6\u306e\u6700\u9069\u5316\u307e\u3068\u3081"},"content":{"rendered":"

\u306f\u3058\u3081\u306b<\/h2>\n

\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u7cbe\u88fd\u306f\u6297\u4f53\u7cbe\u88fd\u3084\u514d\u75ab\u6c88\u964d\u306a\u3069\u3001\u5e83\u304f\u5229\u7528\u3055\u308c\u3066\u3044\u308b\u7cbe\u88fd\u6cd5\u3067\u3059<\/strong>\u3002\u4e00\u822c\u7684\u306b\u306f\u3001PBS, pH 7.2\u306e\u3088\u3046\u306a\u751f\u7406\u7684\u6761\u4ef6\u3067\u306e\u7d50\u5408\u3068\u3001100 mM \u30b0\u30ea\u30b7\u30f3, pH 2.5-3.0\u306e\u3088\u3046\u306a\u9178\u6eb6\u51fa\u3067\u7cbe\u88fd\u3092\u884c\u3044\u307e\u3059\u304c\u3001\u69d8\u3005\u306a\u7d50\u5408\u30fb\u6eb6\u51fa\u6761\u4ef6\u304c\u691c\u8a0e\u3055\u308c\u308b\u3053\u3068\u304c\u3042\u308a\u307e\u3059\u3002\u4eca\u56de\u306f\u3001\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u7cbe\u88fd\u306b\u304a\u3051\u308b\u7d50\u5408\u30fb\u6eb6\u51fa\u6761\u4ef6\u3092\u3054\u7d39\u4ecb\u3057\u307e\u3059<\/strong>\u3002<\/p>\n

\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u7cbe\u88fd\u306b\u304a\u3051\u308b\u7d50\u5408\u6761\u4ef6<\/h2>\n

pH 8.2\u3067IgG\u7d50\u5408\u30ad\u30e3\u30d1\u30b7\u30c6\u30a3\u304c\u6700\u5927\u3068\u306a\u308bProtein A<\/strong>\u306b\u5bfe\u3057\u3066\u3001Protein G\u306fpH\u4f9d\u5b58\u6027\u304c\u4f4e\u304f\u5e83\u7bc4\u56f2\u306epH\u9818\u57df\u3067\u7d50\u5408\u30ad\u30e3\u30d1\u30b7\u30c6\u30a3\u304c\u7dad\u6301\u3055\u308c\u307e\u3059<\/strong>\u3002Protein A\/G\u306fProtein A\u3068G\u306eIgG\u7d50\u5408\u30b5\u30a4\u30c8\u3092\u878d\u5408\u3055\u305b\u305f\u7d44\u307f\u63db\u3048\u30bf\u30f3\u30d1\u30af\u8cea\u3067\u3001Protein G\u540c\u69d8\u306b\u4f4e\u3044pH\u4f9d\u5b58\u6027\u3092\u793a\u3057\u307e\u3059\u3002<\/p>\n

\"optimization-of-affinity-purification-fig\"<\/a>

\u56f3 \u30a2\u30eb\u30d5\u30a1\u30d9\u30c3\u30c8\u30bf\u30f3\u30d1\u30af\u8cea\u306epH\u4f9d\u5b58\u6027<\/p><\/div>\n

\u5869\u306b\u95a2\u3057\u3066\u306f\u3001\u901a\u5e38150 mM NaCl\u306a\u3069\u306e\u5869\u304c\u6dfb\u52a0\u3055\u308c\u307e\u3059\u304c\u3001Protein A\u3067\u306f1 M\u4ee5\u4e0a\u306e\u5869\u306e\u6dfb\u52a0\u306b\u3088\u308a\u30de\u30a6\u30b9IgG1\u306e\u7d50\u5408\u304c\u5897\u5f37\u3055\u308c\u308b\u3053\u3068\u304c\u77e5\u3089\u308c\u3066\u3044\u307e\u3059<\/strong>\u3002Pierce IgG Binding Buffer<\/a>\u306fImmobilized Protein A, G, A\/G\u3092\u4f7f\u7528\u3057\u305f\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u7cbe\u88fd\u306b\u5404\u7d44\u6210\u304c\u6700\u9069\u5316\u3055\u308c\u3066\u304a\u308a\u3001\u4e00\u822c\u7684\u306a\u7d50\u5408\u30d0\u30c3\u30d5\u30a1\u30fc\u306b\u6bd4\u3079\u3066\u7d50\u5408\u30ad\u30e3\u30d1\u30b7\u30c6\u30a3\u306b\u304a\u3051\u308b\u6539\u5584\u304c\u671f\u5f85\u3067\u304d\u307e\u3059<\/strong>\u3002\u8840\u6e05\u3084\u8179\u6c34\u30921:1\u5e0c\u91c8\u3059\u308b\u3060\u3051\u306eready-to-use\u306a\u88fd\u54c1\u3067\u3059\u3002<\/p>\n\n\n\n\n\n\n\n\n\n\n\n
\u88681 IgG Binding Buffer\u3068\u30c8\u30ea\u30b9\u30d0\u30c3\u30d5\u30a1\u30fc\u9593\u306eIgG\u7d50\u5408\u91cf\u306e\u6bd4\u8f03<\/caption>\n
<\/td>\nImmobilized Protein A<\/a><\/td>\nImmobilized Protein G<\/a><\/td>\n<\/tr>\n
\u7d50\u5408\u30d0\u30c3\u30d5\u30a1\u30fc<\/td>\n0.1M\u30c8\u30ea\u30b9\u5869\u9178,
\npH 8.0<\/td>\n		Protein A IgG<\/a>
\nBinding Buffer<\/a><\/td>\n		0.1M\u30c8\u30ea\u30b9\u5869\u9178,
\npH 8.0<\/td>\n		Protein G IgG<\/a>
\nBinding Buffer<\/a><\/td>\n<\/tr>\n
\u30d2\u30c8<\/td>\n19.88<\/td>\n\u00a025.53<\/td>\n\u00a011.68<\/td>\n\u00a023.59<\/td>\n<\/tr>\n
\u30de\u30a6\u30b9<\/td>\n5.25<\/td>\n\u00a07.15<\/td>\n\u00a05.65<\/td>\n\u00a015.05<\/td>\n<\/tr>\n
\u30e9\u30c3\u30c8<\/td>\n4.99<\/td>\n\u00a08.30<\/td>\n\u00a08.43<\/td>\n\u00a011.80<\/td>\n<\/tr>\n
\u30a6\u30b5\u30ae<\/td>\n17.81<\/td>\n\u00a033.19<\/td>\n\u00a021.51<\/td>\n\u00a027.75<\/td>\n<\/tr>\n
\u30d2\u30c4\u30b8<\/td>\n2.15<\/td>\n\u00a010.64<\/td>\n\u00a025.53<\/td>\n\u00a033.33<\/td>\n<\/tr>\n
\u30a6\u30de<\/td>\n6.25<\/td>\n\u00a016.50<\/td>\n\u00a039.19<\/td>\n\u00a021.46<\/td>\n<\/tr>\n
\u30a6\u30b7<\/td>\n6.16<\/td>\n\u00a022.76<\/td>\n\u00a031.72<\/td>\n\u00a048.10<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

\u203b\u6570\u5b57\u306f2mL\u30ec\u30b8\u30f3\u3042\u305f\u308a\u306eIgG\u7d50\u5408\u30ad\u30e3\u30d1\u30b7\u30c6\u30a3(\u5358\u4f4d; mg)<\/p>\n

\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u7cbe\u88fd\u306b\u304a\u3051\u308b\u6eb6\u51fa\u6761\u4ef6<\/h2>\n

\u9178\u6eb6\u51fa\u3055\u308c\u305f\u6297\u4f53\u306f\u3001\u6297\u4f53\u6d3b\u6027\u3092\u7dad\u6301\u3059\u308b\u305f\u3081\u3001\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u7cbe\u88fd\u5f8c\u306f\u901f\u3084\u304b\u306b\u4e2d\u548c\u3055\u308c\u307e\u3059\u3002\u305f\u3060\u3057\u3001\u9178\u6eb6\u51fa\u3067\u4e0d\u53ef\u9006\u306b\u5909\u6027\u3059\u308b\u6297\u4f53\u3082\u3042\u308a\u3001\u305d\u306e\u3088\u3046\u306a\u5834\u5408\u306b\u306f\u7570\u306a\u308b\u6eb6\u51fa\u6761\u4ef6\u304c\u691c\u8a0e\u3055\u308c\u307e\u3059\u3002Gentle Ag\/Ab Binding\/Elution Buffer<\/a>\u306f\u3001\u4e2d\u6027\u4ed8\u8fd1\u306epH (\u7d50\u5408pH\u306f8.0, \u6eb6\u51fapH\u306f6.6)\u3067\u7d50\u5408\u304a\u3088\u3073\u6eb6\u51fa\u3092\u884c\u3046\u7cbe\u88fd\u30b7\u30b9\u30c6\u30e0\u3067\u3001\u6eb6\u51fa\u306f\u9ad8\u6fc3\u5ea6\u306e\u5869\u306b\u3088\u308a\u884c\u3044\u307e\u3059<\/strong>\u3002<\/p>\n

\u9178\u6eb6\u51fa\u3084\u5869(High Salt)\u6eb6\u51fa\u306e\u307b\u304b\u3001\u3055\u307e\u3056\u307e\u306a\u6eb6\u51fa\u6761\u4ef6\u304c\u5831\u544a\u3055\u308c\u3066\u3044\u307e\u3059\u3002\u9178\u6eb6\u51fa\u306f\u6700\u521d\u306b\u691c\u8a0e\u3055\u308c\u308b\u6eb6\u51fa\u6761\u4ef6\u3067\u3059\u304c\u3001\u9178\u6eb6\u51fa\u3067\u52b9\u7387\u7684\u306b\u6eb6\u51fa\u3067\u304d\u306a\u3044\u5834\u5408\u306b\u306f\u30a2\u30eb\u30ab\u30ea\u6eb6\u51fa\u3067\u52b9\u7387\u7684\u306b\u6eb6\u51fa\u3067\u304d\u308b\u3053\u3068\u3082\u3042\u308a\u307e\u3059<\/strong>\u3002\u9178\/\u30a2\u30eb\u30ab\u30ea\u3084\u5869\u6eb6\u51fa\u306e\u307b\u304b\u3001\u3053\u308c\u307e\u3067\u306b\u5831\u544a\u3055\u308c\u3066\u3044\u308b\u6eb6\u51fa\u6761\u4ef6\u3092\u4ee5\u4e0b\u306b\u6dfb\u3048\u307e\u3059\u3002<\/p>\n\n\n\n\n\n\n\n\n\n
\u88682 \u6eb6\u51fa\u6761\u4ef6\u306e\u4f8b<\/caption>\n
\u6eb6\u51fa\u6761\u4ef6<\/td>\n\u4f8b<\/td>\n<\/tr>\n
\u9178<\/td>\nIgG Elution Buffer, pH 2.8<\/a>
\n100 mM \u30b0\u30ea\u30b7\u30f3(\u5869\u9178), pH 2.5-3.0
\n100 mM \u30af\u30a8\u30f3\u9178, pH 3.0<\/td>\n<\/tr>\n
\u30a2\u30eb\u30ab\u30ea<\/td>\n50-100 mM \u30c8\u30ea\u30a8\u30c1\u30eb\u30a2\u30df\u30f3\u307e\u305f\u306f\u30c8\u30ea\u30a8\u30bf\u30ce\u30fc\u30eb\u30a2\u30df\u30f3, pH 11.5
\n150 mM \u6c34\u9178\u5316\u30a2\u30f3\u30e2\u30cb\u30a6\u30e0
\n100 mM \u30b0\u30ea\u30b7\u30f3(\u6c34\u9178\u5316\u30ca\u30c8\u30ea\u30a6\u30e0), pH 10.0<\/td>\n<\/tr>\n
\u5869<\/td>\nGentle Ag\/Ab Elution Buffer, pH 6.6<\/a>
\n5 M \u5869\u5316\u30ea\u30c1\u30a6\u30e0
\n3.5 M \u5869\u5316\u30de\u30b0\u30cd\u30b7\u30a6\u30e0\u307e\u305f\u306f\u5869\u5316\u30ab\u30ea\u30a6\u30e0
\n2.5 M \u30e8\u30a6\u5316\u30ca\u30c8\u30ea\u30a6\u30e0\u307e\u305f\u306f\u30e8\u30a6\u5316\u30ab\u30ea\u30a6\u30e0
\n0.2-3.0 M \u30c1\u30aa\u30b7\u30a2\u30f3\u9178\u30ca\u30c8\u30ea\u30a6\u30e0
\n0.1 M \u30c8\u30ea\u30b9\u9162\u9178, 2.0 M \u5869\u5316\u30ca\u30c8\u30ea\u30a6\u30e0, pH 7.7<\/td>\n<\/tr>\n
\u5909\u6027\u5264<\/td>\n2-6 M \u30b0\u30a2\u30cb\u30b8\u30f3\u5869\u9178
\n2-8 M \u30a6\u30ec\u30a2
\n1.0 M\u30c1\u30aa\u30b7\u30a2\u30f3\u9178\u30a2\u30f3\u30e2\u30cb\u30a6\u30e0
\n1% \u30c7\u30aa\u30ad\u30b7\u30b3\u30fc\u30eb\u9178\u30ca\u30c8\u30ea\u30a6\u30e0
\n1% SDS<\/td>\n<\/tr>\n
\u6709\u6a5f\u6eb6\u5a92<\/td>\n10% \u30b8\u30aa\u30ad\u30b5\u30f3
\n50% \u30a8\u30c1\u30ec\u30f3\u30b0\u30ea\u30b3\u30fc\u30eb, pH 8-11.5<\/td>\n<\/tr>\n
\u7af6\u5408<\/td>\n0.1 M\u4ee5\u4e0a\u306e\u30ea\u30ac\u30f3\u30c9<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

\u304a\u308f\u308a\u306b<\/h2>\n

\u7d50\u5408\u30fb\u6eb6\u51fa\u306e\u6761\u4ef6\u306f\u3001\u7cbe\u88fd\u5f8c\u306e\u6297\u4f53\u3084\u6297\u539f\u306e\u6d3b\u6027\u7dad\u6301\u306e\u307b\u304b\u3001\u53ce\u7387\u3084\u30a2\u30d5\u30a3\u30cb\u30c6\u30a3\u30ab\u30e9\u30e0\u306e\u5bff\u547d(\u518d\u751f\u56de\u6570)\u306b\u3082\u5f71\u97ff\u3059\u308b\u3053\u3068\u304c\u3042\u308a\u307e\u3059\u3002<\/p>\n

\u3010\u7121\u6599\u30c0\u30a6\u30f3\u30ed\u30fc\u30c9\u3011\u30bf\u30f3\u30d1\u30af\u8cea\u89e3\u6790\u30ef\u30fc\u30af\u30d5\u30ed\u30fc\u30cf\u30f3\u30c9\u30d6\u30c3\u30af<\/h3>\n

\u52b9\u7387\u7684\u306a\u30bf\u30f3\u30d1\u30af\u8cea\u306e\u62bd\u51fa\u304b\u3089\u30a6\u30a7\u30b9\u30bf\u30f3\u30d6\u30ed\u30c3\u30c6\u30a3\u30f3\u30b0\u306e\u89e3\u6790\u30c4\u30fc\u30eb\u307e\u3067\u3001\u5305\u62ec\u7684\u306b\u30bd\u30ea\u30e5\u30fc\u30b7\u30e7\u30f3\u3092\u7d39\u4ecb\u3057\u3066\u304a\u308a\u307e\u3059\u3002PDF\u30d5\u30a1\u30a4\u30eb\u306e\u30c0\u30a6\u30f3\u30ed\u30fc\u30c9\u3092\u3054\u5e0c\u671b\u306e\u65b9\u306f\u3001\u4e0b\u8a18\u30dc\u30bf\u30f3\u3088\u308a\u304a\u7533\u3057\u8fbc\u307f\u304f\u3060\u3055\u3044\u3002<\/p>\n

\u30c0\u30a6\u30f3\u30ed\u30fc\u30c9\u3059\u308b<\/a><\/p>\n

 <\/p>\n

\u7814\u7a76\u7528\u306b\u306e\u307f\u4f7f\u7528\u3067\u304d\u307e\u3059\u3002\u8a3a\u65ad\u76ee\u7684\u304a\u3088\u3073\u305d\u306e\u624b\u7d9a\u304d\u4e0a\u3067\u306e\u4f7f\u7528\u306f\u3067\u304d\u307e\u305b\u3093\u3002<\/p>\n","protected":false},"excerpt":{"rendered":"

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