A CE-LIF method to monitor autophagy by directly detecting LC3 proteins in HeLa cells.
A fast, simple and quantitative approach was established for monitoring autophagy in HeLa cells by directly detecting the conversion of green fluorescent protein (GFP) labelled autophagy markers, GFP-LC3-I to GFP-LC3-II, in crude cellular extract using capillary electrophoresis (CE) with laser-induced fluorescence (LIF). Compared with the traditional methods, this proposed method is simpler and more reliable. Moreover, high sensitivity, with a limit of detection (LOD) of 0.48 ppt (bovine serum albumin as standard protein), was obtained by an in-capillary derivatization method coupled with a field-enhanced sample injection (FESI) technique, this may allow the success of this technique in the detection of endogenous markers of autophagy in cells.
Beijing National Laboratory for Molecular Sciences, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, PR China.
Yue
Y
Jin
Cheng
C
Chen
Lingchen
L
Meng
Jitao
J
Chen
Meixian
M
Li
Zhiwei
Z
Zhu
Jian
J
Lin
D001343:Autophagy; D019075:Electrophoresis, Capillary / Q000379:methods*; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007834:Lasers; D057230:Limit of Detection; D008869:Microtubule-Associated Proteins / Q000032:analysis*
C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; D049452:Green Fluorescent Proteins