RIPA裂解液(RIPA的本意是Radio Immunoprecipitation Assay)是一种传统的细胞组织快速裂解液，主要用于从动物细胞和组织中提取可溶性蛋白，其裂解得到的蛋白样品可以用于常规的Western Blot、IP及Elisa等实验。RIPA裂解液的配方有很多种，根据其裂解强度大致可以分为强、中、弱三类。用RIPA裂解液裂解得到的蛋白样品，可以使用 BCA蛋白定量试剂盒 (货号：ZJ101或ZJ102)测定蛋白浓度。
取适当量的裂解液(每1×106个细胞需要约50~100μL或每20mg组织样本需要约150~250μL)，在使用前数分钟内将 蛋白酶抑制剂 按1:100(V/V)加入其中( 蛋白酶抑制剂 需另行购买，货号：GRF101)；
注意：如所需提取的为磷酸化蛋白，还需在裂解液中按1:100(V/V)加入 磷酸酶抑制剂 (货号：GRF102)。
样本裂解(需在冰上操作)：
◆ 对于贴壁细胞：
① 弃去培养基，用1×PBS(货号：PS110，或生理盐水、无血清培养液)洗一遍(如果血清中的蛋白对实验没有干扰，也可以不洗)；
② 尽可能地弃去PBS(多余的液体将降低裂解液的浓度)；
③ 按照每1×106个细胞需要约50~100μL的比例加入RIPA裂解液，比如6孔板每孔细胞量大约需加入150~250μL裂解液。用移液器吹打数下，使裂解液和细胞充分接触。通常裂解液接触细胞1~2s后，细胞就会被裂解；
④ 将所有液体转入新的离心管中；
◆ 对于悬浮细胞：
① 将细胞转移至离心管中，离心收集细胞，弃去培养基；
② 用1×PBS(货号：PS110，或生理盐水、无血清培养液)洗一遍(如果血清中的蛋白对实验没有干扰，也可以不洗)；
③ 尽可能地弃去PBS(多余的液体将降低裂解液的浓度)；
④ 按照每1×106个细胞需要约50~100μL的比例加入RIPA裂解液，比如6孔板每孔细胞量大约需加入150~250μL裂解液。用移液器吹打数下，使裂解液和细胞充分接触。充分裂解后应没有明显的细胞沉淀。如果细胞量较多，必须分装成5×105~1×106个细胞/管，然后再裂解；
◆ 对于组织样品：
① 把组织剪切成细小的碎片；
② 按照每20mg组织样本加入150~250μL裂解液的比例加入裂解液；
注意：如果样本裂解不充分，可以适当提高裂解液的用量；若需要高浓度的蛋白样品，也可适当降低裂解液的用量。
③ 用玻璃匀浆器匀浆，直至样本充分裂解；
注意：若组织样本非常细小，可以适当剪切后直接加入裂解液，通过强烈涡旋振荡使其裂解充分。
充分裂解后，10,000~14,000×g离心3~5min，小心地将上清液(蛋白样品)移入新的离心管中，即可进行后续的PAGE凝胶电泳、Western Blot和免疫沉淀等操作。得到的蛋白样品可分装并长期保存于-80℃。
1. 4℃保存时，裂解液中的SDS易析出，使用前请置于37℃使其完全溶解，待恢复到室温即可使用；
2. 裂解样品的所有步骤都需在冰上或4℃进行；
3. RIPA裂解液的裂解产物中经常会出现一小团透明胶状物，此为正常现象，该物质为基因组DNA等形成的复合物。如不检测和基因组DNA紧密结合的蛋白，可以直接离心取上清用于后续实验；若需要检测此类蛋白，则可以通过超声处理打散该透明胶状物，随后离心取上清即可用于后续实验。但如果检测一些常见的转录因子，例如NF-kappaB、p53等，通常不必进行超声处理就可完成检测；
4. 为了您的安全和健康，请穿实验服并戴一次性手套操作；
5. 本产品仅限科研使用。
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