使用说明：
1. 准备试剂：室温融解并混匀膜蛋白抽提试剂A和B，融解后立即置于冰浴上。取适量的膜蛋白抽提试
    剂A和B备用，在使用前数分钟内加入PMSF，使PMSF的最终浓度为1mM。
2. 准备细胞或组织样品：
    a. 对于细胞
    (1) 收集细胞
    对于贴壁细胞：培养约2000-5000万细胞，用PBS洗一遍，用细胞刮子刮下细胞或用含有EDTA但不含
    胰酶的细胞消化液处理细胞使细胞不再贴壁很紧，并用移液器吹打下细胞。离心收集细胞，吸除上
    清，留下细胞沉淀备用。尽量避免用胰酶消化细胞，以免胰酶降解需抽提的目的膜蛋白。
    对于悬浮细胞：培养约2000-5000万细胞，直接离心收集细胞，吸除上清，留下细胞沉淀备用。
    (2) 洗涤细胞：用适量冰浴预冷的PBS轻轻重悬细胞沉淀，取少量细胞用于计数，剩余细胞4℃，
    600g离心5分钟沉淀细胞。弃上清，随后4℃，600g离心1分钟，以沉淀离心管管壁上的残留液体并进
    一步沉淀细胞，尽最大努力吸尽残留液体。
    (3) 细胞预处理：把1毫升临用前添加了PMSF的膜蛋白抽提试剂A加入至2000-5000万细胞中，轻轻
    并充分悬浮细胞，冰浴放置10-15分钟。
    b. 对于组织：取约100毫克组织，用剪刀尽量小心剪切成细小的组织碎片。加入1毫升临用前添加
    了PMSF的膜蛋白抽提试剂A，轻轻悬浮组织碎片，冰浴放置10-15分钟。注：如果组织样品比较少，
    也可以使用更少的组织量，例如30-50mg，后续试剂的用量及操作步骤不变；组织用量较少时，最后
    获得的膜蛋白也较少。
3. 细胞或组织样品的破碎及破碎效果的鉴定：把细胞悬液或组织样品转移到一适当大小的冰浴预冷
    玻璃匀浆器中，匀浆约30-50下。匀浆效果与细胞类型和组织类型相关，不同细胞或组织所需的匀浆
    次数有所不同，需自行优化。通常可以在匀浆30次后取约2-3微升细胞或组织匀浆液滴在盖玻片上并
    在显微镜下观察，如见细胞核周晕环(A shiny ring around the nuclei)或完整的细胞形态，说明
    细胞仍完整。如果有70-80%的细胞均无核周晕环和完整细胞形态，说明细胞已经充分破碎，则进行
    下一步实验。否则，重新匀浆10-30次直到细胞至少70%已经破碎。同时记录对于该细胞的匀浆次
    数，通常在后续实验时不必再摸索匀浆次数。另外需注意特定的匀浆次数和匀浆器也有关，需同时
    注意记录使用的是哪一个匀浆器。
    注：如果没有适当的玻璃匀浆器，对于培养的细胞也可以采用冻融法来破碎细胞。把步骤2中的样
    品在液氮和室温依次反复冻融两次，然后取少量样品在显微镜下检测细胞破碎的程度。如果细胞破
    碎的程度不足70%，可增加冻融次数，直到细胞破碎的程度大于70%。
4. 去除细胞核和未破碎的细胞： 4℃，700g离心10分钟，小心收集上清液至一新的离心管中。吸取
    上清时切勿接触沉淀！可以有约30-50微升上清液残留不予吸取，以保证吸取的上清液有较高的纯
    度。
5. 沉淀细胞膜碎片：4℃，14000g离心30分钟，以沉淀细胞膜碎片。
6. 收集细胞浆蛋白：吸取上清即为细胞浆蛋白，可-70℃保存备用。吸取上清时可以有30-50微升上清
    残留，以避免接触沉淀导致上清样品被污染。每5000万细胞使用本产品裂解可获得5-30mg细胞浆蛋
    白, 不同细胞有所不同。
7. 抽提膜蛋白：4℃，14000g离心10秒，尽最大努力吸尽上清。可以轻轻触碰到沉淀，甚至吸走很少
    量的沉淀。加入膜蛋白抽提试剂B 200微升(如有必要，也可以加大到300微升)，最高速剧烈Vortex
    5秒重悬沉淀，冰浴5-10分钟。重复前述步骤的vortex和冰浴孵育1-2次，以充分抽提膜蛋白。随
    后，4℃，14000g离心5分钟，收集上清即为细胞膜蛋白溶液。可-70℃保存备用。对于一些有特殊用
    途的膜蛋白，可自行配制适当的膜蛋白抽提试剂进行膜蛋白抽提。每5000万细胞使用本产品裂解可
    获得0.3-3mg细胞膜蛋白, 不同细胞有所不同。

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