[交流] 【求助/交流】求教做免疫共沉淀的人

我没做过免疫共沉淀，现在要设计免疫共沉淀实验，为了更高的成功率和实验质量，现在请做过的高手简要介绍一下免疫共沉淀最可能遇到的几种问题

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1楼 2010-12-10 16:37:12

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上海生工有试剂盒卖，很好用！
Immunoprecipitation Kit
Components BS688 For 20 reactions BS689 For 20 reactions BS690 For 20 reactions
10x Lysis buffer 10 ml 10 ml 10 ml
PMSF 0.6ml 0.6ml 0.6ml
10x IP 50 ml 50 ml 50 ml
SDS 5 ml 5 ml 5 ml
NaCl 10 ml 10 ml 10 ml
Protein A-Agarose 0.4 ml / /
Protein A/G Plus-Agarose / 0.4 ml /
Protein A-Sepharose / / 0.6 ml
20x PBS 20 ml 20 ml 20 ml
1x SDS gel-loading buffer 1 ml 1ml 1 ml
Spin columns and tubes 20 20 20
Price ¥1,520 ¥2,250 ¥1,920
Description:
Immunoprecipitation (IP) is a method by which a protein can be specifically purified from a complex mixture of proteins using a specific antibody and a matrix that binds the antibody. The matrix bound protein (via the specific antibody) can then be separated from the mixture by centrifugation. The matrices commonly used are agarose or sepharose bound Protein A or protein A/G PlUS. IP is one of the most widely used immunochemical techniques. IP followed by SDS-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and immunoblotting is routinely used in a variety of applications: to measure the molecular weights of protein antigens, study protein/protein interactions, determine specific enzymatic activity, monitor protein post-translational modifications and determine the presence and quantity of proteins. The IP technique also enables the detection of rare proteins, which otherwise would be difficult to detect, since immunoprecipitation can concentrate them as much as 10,000-fold..
Applications:The Protein-A Immunoprecipitation Kit is especially designed to allow maximal recovery of immunoprecipitates. The whole process is performed in mini-spin columns, instead of in microcentrifuge tubes, which enables convenient washing of the antigen-antibody-bound beads.
Features:Easy access.;Minimal loss of antigen-antibody bound beads during washing.；Minimal non-specific signals by increasing the stringency of the washing steps.
Storage Condition:Stored at 4℃.and PMSF should be stored at -20℃.