TABLE 1.
Fluorescence-activated cell sorting analysis of CaV1.2 ± auxiliary subunits
FACS results obtained after the transient transfection of CaV1.2-HA wt in either HEKT control cells, stable CaVβ3 cells, or stable CaVα2bδ cells. One day (24 h) after transfection, the cells were incubated with anti-HA FITC conjugate (10 μg/ml) at room temperature for 45 min. FACS separation of FITC-positive cells was performed on a FACScalibur® flow cytometer (Becton Dickinson), and fluorescence was quantified using CellQuest software (Becton Dickinson). The results are reported as percentage values of cells in M2. The data were pooled from experiments carried out over a period of 8 months. The data are shown as the means ± S.E. of the individual experiments, and the number of experiments appears in parentheses. ND, not determined.
| Construct transient expression | Cell lines |
|||
|---|---|---|---|---|
| HEKT | HEKT CaVβ3 stable | HEKT CaVα2bδ stable | HEKT CaVβ3 + CaVα2bδ stable | |
| % | % | % | % | |
| Cells without antibody | 0.3 ± 0.1 (32) | 0.18 ± 0.04 (20) | 0.54 ± 0.07 (3) | 2.8 ± 0.2 (3) |
| Cells with antibody | 1.1 ± 0.3 (19) | 1.0 ± 0.4 (19) | 0.93 ± 0.05 (3) | 4.2 ± 0.4 (3) |
| CaV1.2-HA wt | 4.5 ± 0.5 (25) | 23 ± 1 (29) | 6 ± 1 (3) | 19 ± 2 (3) |
| CaV1.2-HA wt + CaVβ3 | 25 ± 2 (3) | ND | 21 ± 2 (3) | ND |
| CaV1.2-HA wt + CaVβ2a | 14 ± 1 (3) | ND | ND | ND |
| CaV1.2-HA wt + CaVβ4 | 22 ± 3 (3) | ND | ND | ND |
| CaV1.2-HA wt + CaVα2bδ + CaM wt | 2 ± 1 (3) | ND | ND | ND |
| CaV1.2-HA wt + CaVα2bδ | 8 ± 3 (5) | 26 ± 1 (3) | ND | ND |
| CaV1.2-HA wt + CaM wt | 1.5 ± 0.3 (3) | 19.1 ± 0.3 (3) | 2.7 ± 0.4 (3) | ND |
| CaV1.2-HA wt + CaM1,2,3,4 | 1.4 ± 0.1 (3) | 19.1 ± 0.6 (3) | ND | ND |
| CaV1.2-HA wt + CaVβ3 + CaVα2bδ | 23 ± 2 (5) | ND | ND | ND |