Biomedical Engineering Department, Texas A&M University, College Station, TX 77843, USA.
Electrophoresis. 2011 Jun;32(12):1433-7. doi: 10.1002/elps.201000684. Epub 2011 May 30.
IEF protein binary separations were performed in a 12-μL drop suspended between two palladium electrodes, using pH gradients created by electrolysis of simple buffers at low voltages (1.5-5 V). The dynamics of pH gradient formation and protein separation were investigated by computer simulation and experimentally via digital video microscope imaging in the presence and absence of pH indicator solution. Albumin, ferritin, myoglobin, and cytochrome c were used as model proteins. A drop containing 2.4 μg of each protein was applied, electrophoresed, and allowed to evaporate until it splits to produce two fractions that were recovered by rinsing the electrodes with a few microliters of buffer. Analysis by gel electrophoresis revealed that anode and cathode fractions were depleted from high pI and low pI proteins, respectively, whereas proteins with intermediate pI values were recovered in both fractions. Comparable data were obtained with diluted bovine serum that was fortified with myoglobin and cytochrome c.
IEF 蛋白质双相分离在 12μL 液滴中进行，该液滴悬挂在两个钯电极之间，使用低电压（1.5-5V）电解简单缓冲液产生的 pH 梯度。通过计算机模拟和数字视频显微镜成像实验研究了 pH 梯度形成和蛋白质分离的动力学，实验中存在和不存在 pH 指示剂溶液。白蛋白、铁蛋白、肌红蛋白和细胞色素 c 被用作模型蛋白。将含有每种蛋白质 2.4μg 的液滴施加、电泳并允许蒸发，直到它分裂产生两个分数，然后用几微升缓冲液冲洗电极来回收这些分数。凝胶电泳分析表明，阳极和阴极部分分别耗尽了高 pI 和低 pI 蛋白，而具有中等 pI 值的蛋白质则在两个部分中都得到了回收。用含有肌红蛋白和细胞色素 c 的稀释牛血清进行了类似的实验，得到了可比的数据。
