| Cell Line Species/Tissue: |
Human
/
umbilical vein endothelial cells |
| Transfection Reagent: |
HiPerFect |
Nucleic Acid:
|
siRNA (dsRNA) |
Growth Medium:
|
PromoCell Endothelial cell basal medium |
Percent Serum (%):
|
2 % |
Reporter System:
|
|
Plasmid Purification Method:
|
|
Plate Format:
|
6-well plate |
Number of Cells:
|
100.000 and 500.000 |
Percent Confluence(%):
|
30-80% (approx) |
Amount of Nucleic Acid (µg):
|
0.3 - 1.8 µg |
Amount of Enhancer (µl):
|
|
Amount of Reagent (µl):
|
12 µl |
Complex Incubation on Cells (hrs):
|
24 h |
Analysis Performed Post-Transfection (hrs):
|
48 h |
Transfection Efficiency (%):
|
more than 93 % (measured by FACS) |
| Knockdown Efficiency (%): |
more than 93 % (measured by FACS) |
| |
| Any modifications to the protocol?:
|
| See notes |
| Notes: |
| I performed the transfection in specialized medium (Endothelial cell basal medium from Promocell, containing 2 %FCS), and repeated transfection after 24h. Western Blot was used for kockdown analysis.
It works nicely on HUVECs with higher efficiency than Oligofectamine or RNAiFect and shows no obvious alteration in cell morphology (like with RNAiFect) or cytotoxic effects (Oligofectamine).
siRNA used: connexin 43; only negative control siRNA from Qiagen was Alexa 488 labelled |
| References: |
| |