The cells were plated on 96-well plates for On-Cell Western assays or on 12-mm coverslips in 6-well plates for electrophysiological experiments, maintained in DMEM containing 10% fetal bovine serum (both Life Technologies), and incubated at 37 °C in 5% CO2. The cells were transfected 24 h after plating with the BK channel α-subunit or co-expressed with β1-subunit in a 2:1 ratio of cDNA using Lipofectamine 2000 (Thermo Fisher) or Polyjet (tebu-bio) unless otherwise specified. All expression constructs were previously described (12 (link), 34 (link)). The cells were used for experiments 24–48 h post-transfection.
Transient Transfection of HEK293 Cells
The cells were plated on 96-well plates for On-Cell Western assays or on 12-mm coverslips in 6-well plates for electrophysiological experiments, maintained in DMEM containing 10% fetal bovine serum (both Life Technologies), and incubated at 37 °C in 5% CO2. The cells were transfected 24 h after plating with the BK channel α-subunit or co-expressed with β1-subunit in a 2:1 ratio of cDNA using Lipofectamine 2000 (Thermo Fisher) or Polyjet (tebu-bio) unless otherwise specified. All expression constructs were previously described (12 (link), 34 (link)). The cells were used for experiments 24–48 h post-transfection.
Corresponding Organization :
Other organizations : University of Edinburgh
Protocol cited in 1 other protocol
Variable analysis
- Transfection of BK channel α-subunit or co-expressed with β1-subunit in a 2:1 ratio of cDNA
- Expression of BK α- or β1-subunit as determined by mRNA, protein, or functional assays
- HEK 293 cells between passage 18 and 30, originally obtained from ATCC
- HEK 293 cells do not express endogenous the BK α- or β1-subunit
- HEK 293 cells were cultured and maintained in DMEM containing 10% fetal bovine serum at 37 °C in 5% CO2
- Transfection was performed 24 h after plating the cells
- Cells were used for experiments 24–48 h post-transfection
- Not explicitly mentioned
- Not explicitly mentioned
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