Janet Lynch1, JiWoo Chung1, Zhen Huang1, Vincen Pierce1, Noah S Saunders1, Li Niu2. 1. Department of Chemistry and Center for Neuroscience Research, University at Albany, State University of New York, Albany, NY, 12222, United States. 2. Department of Chemistry and Center for Neuroscience Research, University at Albany, State University of New York, Albany, NY, 12222, United States. Electronic address: lniu@albany.edu.
Abstract
BACKGROUND: Transient expression of proteins in mammalian cells is a key technique for many functional and structural studies of human and higher eukaryotic genes as well as for the production of recombinant protein therapeutics. Maximizing the expression efficiency to achieve a higher expression yield is desirable and may be even critical when, for instance, an expressed protein must be characterized at the single-cell level. NEW METHODS: Our goal was to develop a simple method by which protein expression yield in human embryonic kidney (HEK)-293 cells could be enhanced with a brief treatment of dimethyl sulfoxide (DMSO) solution. RESULTS: By expressing green fluorescent protein (GFP) as a reporter protein using the calcium phosphate transfection method and imaging a large population of cells, we found that a 5-min exposure of 10 % DMSO to HEK-293 cells, 4 h after transfection of the protein of interest, leads to ∼1.6-fold increase in the expression yield without causing any appreciable cytotoxicity. By expressing an α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and separately a kainate receptor in HEK-293 cells and measuring glutamate-induced whole-cell current response, we also found that such a brief DMSO treatment did not affect channel activity. CONCLUSION: This method is simple, efficient and inexpensive to use for enhancing transient transfection yield in HEK-293 cells.
BACKGROUND: Transient expression of proteins in mammalian cells is a key technique for many functional and structural studies of human and higher eukaryotic genes as well as for the production of recombinant protein therapeutics. Maximizing the expression efficiency to achieve a higher expression yield is desirable and may be even critical when, for instance, an expressed protein must be characterized at the single-cell level. NEW METHODS: Our goal was to develop a simple method by which protein expression yield in human embryonic kidney (HEK)-293 cells could be enhanced with a brief treatment of dimethyl sulfoxide (DMSO) solution. RESULTS: By expressing green fluorescent protein (GFP) as a reporter protein using the calcium phosphate transfection method and imaging a large population of cells, we found that a 5-min exposure of 10 % DMSO to HEK-293 cells, 4 h after transfection of the protein of interest, leads to ∼1.6-fold increase in the expression yield without causing any appreciable cytotoxicity. By expressing an α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and separately a kainate receptor in HEK-293 cells and measuring glutamate-induced whole-cell current response, we also found that such a brief DMSO treatment did not affect channel activity. CONCLUSION: This method is simple, efficient and inexpensive to use for enhancing transient transfection yield in HEK-293 cells.
Authors: Maooz Awan; Iryna Buriak; Roland Fleck; Barry Fuller; Anatoliy Goltsev; Julie Kerby; Mark Lowdell; Pavel Mericka; Alexander Petrenko; Yuri Petrenko; Olena Rogulska; Alexandra Stolzing; Glyn N Stacey Journal: Regen Med Date: 2020-04-28 Impact factor: 3.806
Authors: Joana Galvao; Benjamin Davis; Mark Tilley; Eduardo Normando; Michael R Duchen; M Francesca Cordeiro Journal: FASEB J Date: 2013-12-10 Impact factor: 5.191