AKTA--实验室层析柱装柱技术介绍PPT参考课件.ppt

* * * * * * * * * * * * 实验室层析柱装柱技术介绍 * Column packing 好的柱效是层析成功的关键 不同种类的凝胶、不同的层析柱有不同的装柱方法 柱效测定的方法 层析柱的维护 * Column packing Rabies vaccine Sepharose 4 Fast Flow 5% column vol sample * Column packing ml mAU*ml (volume) % mAU No Ret Area Area/Peak area Height 1 36.27 27.8394 84.45 8.218 2 118.24 5.1267 15.55 0.649 每米塔板数：N=1600 ml mAU*ml (volume) % mAU No Ret Area Area/Peak area Height 1 36.14 50.6325 98.48 16.974 2 118.54 0.7791 1.52 0.131 每米塔板数：N=3500 * Column packing N/m= 5.54 (Ve/Vh)2100/L As = b/a * Column packing 检测装柱效果一般用0.5% (30um介质)或1% (90um介质) 柱体积的1%丙酮测柱效和峰形 The linear flow rate should be 30 cm/h for 34 μm and 50 μm media and 20 cm/h for 90 μm media * * 线性流速 对体积不同的柱子来说，体积流速不能直接用作比较， 一般都换算成线性流速，再作比较： 体积流速 (ml/min) x 60 线性流速 (cm/hr) = 揪揪揪揪揪揪 柱子横切面积 (cm2) Column packing * Column packing It is important that the column is properly equilibrated before the packing is evaluated (~ 2 column volumes). It is preferable to run three test runs to see whether the test values are stable. If a poor result improves during this test, the reason can be that the column was not properly equilibrated. To check that the bed is stable, run the column at 70% packing pressure for 20 hours and test it again (preferably three test runs). Note that pressure spikes may cause poor packing (cracking). If this happens, an air trap and a pressure relief valve are needed between the pump and the column. The pressure relief valve should be placed between the air trap and column. * Column packing * 装柱注意点 选择一根设计良好的柱子 仔细混合均匀凝胶浆 在生产时使用柱子的温度下装柱 用平缓的动作将凝胶浆一次性加入柱子中 用恒压或恒流装柱 稳定和平衡凝胶柱床 检查柱效，必要时