基于过程优化探索蛋白印迹技术的实验教学设计  

Experimental Teaching Design of Western Blotting Based on Process Optimization

作　　者：葛家祥 尚佳琪 刘江 邓仕婧 林牒 杨琳[2] GE Jiaxiang;SHANG Jiaqi;LIU Jiang;DENG Shijing;LIN Die;YANG Lin(College of Integrated Chinese and Western Medicine,Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China;College of Medical Technology,Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China)

机构地区：[1]天津中医药大学中西医结合学院,天津301617 [2]天津中医药大学医学技术学院,天津301617

出　　处：《实验室研究与探索》2023年第1期223-228,共6页Research and Exploration In Laboratory

基　　金：国家自然科学基金青年项目(81704004);国家自然科学基金面上项目(82174470);全国中医药高等教育“十四五”规划2021年度教育科研课题(YB-20-17);天津市普通高等学校本科教学质量与教学改革研究计划项目(B201006301)。

摘　　要：为了提高蛋白印迹实验的成功率及缩短蛋白印迹实验的周期,探索符合实验教学的过程方案,以蛋白印迹法实验操作为模板,以β-actin(48 kDa)为目的蛋白,以GAPDH(36 kDa)为标准蛋白,通过改变实验条件,计算β-actin与GAPDH显影后灰度值的比值,进行实验过程的优化。并对优化前后实验过程中在制胶、加样、电泳、转膜、封闭以及孵育等环节的成功率、离散度、所用时间等方面进行比较。结果表明,优化后的免疫印迹法能有效地提高成功率、降低离散度、缩短实验时间,β-actin与GAPDH显影后灰度值比值更接近1,达到实验教学的要求。In order to improve the success rate of Western blot experiment and shorten the cycle of Western blot experiment,this paper explores the process scheme in line with experimental teaching The Western blot is used as the template,β-actin(48 kDa)as the target protein and GAPDH(36 kDa)as the standard protein.The ratio ofβ-actin to GAPDH is used to optimize the experimental process.Comparing before and after the optimization of the experimental process,we can see that the success rate is high,the dispersion is low and the time is shortened after the optimization in the links of glue preparation,sample addition,electrophoresis,membrane transfer,sealing and incubation.We conclude that the improved immunoblotting method can effectively improve the success rate,shorten the experimental time and shorten the experimental time.The ratio ofβ-actin to GAPDH is closer to 1,which meets the requirements of experimental teaching.

关 键 词：蛋白印迹 过程优化 方法改进 实验教学 

分 类 号：G642[文化科学—高等教育学] N33[文化科学—教育学] N45[自然科学总论]