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Native gel electrophoresis

XJ Xuechao Jia
CH Chuntian Huang
YH Yamei Hu
QW Qiong Wu
FL Fangfang Liu
WN Wenna Nie
HC Hanyong Chen
XL Xiang Li
ZD Zigang Dong
KL Kangdong Liu
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The Native gel electrophoresis assay was conducted based on the Gel System protocol of the NativePAGE™ Novex® Bis-Tris. Cells were incubated for 30 min on ice in 1× sample buffer (50 mM Bis-Tris, 0.1% n-dodecyl-β-D-maltoside, 6 N HCl, 50 mM NaCl, 0.001% Ponceau S, 10% Glycerol, pH 7.2). After 30 min 20,000 g centrifugation, the supernatant was collected, and the concentration was measured using BCA kit. NativePAGE™ gel (4–16%) electrophoresis was performed at 150 V constant voltages. After running for 30 min the Cathode Buffer-Dark Blue was changed with Light Blue and electrophoresis was resumed for another 80 min. The gel was then transferred in 1 × NuPAGE® Transfer Buffer at 100 mA constant for 1 h and the membrane was fixed with 20 mL of 8% acetic acid for 15 min and immunodetection was directly performed.

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