知识_抗体效价的elisa测定.pptVIP

酶标板的清洗：采用ELx50TM型微孔板洗板机，注射泵驱动的液体输送，条状单排清洗或全板清洗 * 了解转膜电泳原理的应用及操作 Western Blotting or immunoblotting a specific primary antibody 1979, Towbin 1975, Southern, Southern blotting 1977, Alwine, Northern blotting 1979, Towbin, Western Blotting 1982, Reihart, Eastern blotting ,双向蛋白质印记 * * * * Western blotting: Antibodies can be used to detect specific proteins * Alternate detection method – Secondary antibodies * Memebrane: 吸附生命大分子的固体材料 NC, nitrocellulose 硝酸纤维素膜 0.45um 结合率：80 ug Pr/cm2 便宜，可简单快速封闭非特异性抗体的结合 NDM, nylon-densed membrane 尼龙膜 结合率：480 ug Pr/cm2 DBM, diazobenzyloxymethyl 重氮苄氧甲基膜 * * 湿法转移蛋白质：将gel 和membrane 夹在blotter paper 中，浸在转移装置的buffer中，通电45min 或over night 可完成。 Transfer Buffer (500 ml, pH 8.3) glycine 1.450 g (39 mM) Tris base 2.900 g (48 mM) SDS 0.185 g (0.037%) * Using the Semi-Dry Transfer Unit Remove the stacking gel from the gel. 去除堆积凝胶的凝胶。 2.Cut pieces of blotter paper to the size of the gel. note: the blotter paper (and the membrane) not be larger than the gel. Larger pieces may make contact around the gel, and allow the current an alternate route, thereby making transfer inefficient. 裁片的记事簿纸张的大小的凝胶。 注：本记事簿纸（和膜）不大于凝胶。大块的可能使接触周围的凝胶，并允许目前的替 代路线，从而使传输效率。 3. Wearing gloves rinsed of powder, cut a piece of membrane to the size of the gel. Mark the lower right-hand corner to allow for orientation. Pre-wet the membrane in transfer buffer for 2 to 5 minutes. 戴手套清洗粉，剪下一块膜大小的凝胶。标记右下角以便方向。湿膜转移缓冲液为2到5 分钟。 * 4. sandwich blotter paper – membrane- gel- blotter paper make sure no air bubbles are trapped The membrane must therefore be positioned correctly the first time. Do not try to adjust. 三明治 吸墨纸纸–膜凝胶-记事簿纸 确保没有气泡被困 膜因此必须正确定位第一时间。不要试图调整。 * 5. Hold the cover level and slide it gently down onto the stack. 持有的封面和滑动它轻轻地放到堆栈 6. weigh down the lid in order to ensure even contact with the stack.权衡下来的盖子，以确保即使接触堆栈。 7. Connect the unit to