ON-TARGETplus Mouse Cdh5 (12562) siRNA - SMARTpool

Protocol tips

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	HUVECs and MS-1 cells were incubated with 40 nM ON-TARGETplus SMARTpool duplex siRNA targeting Cdh5 (Dharmacon, #L003641; L-041968) or 20 nM siRNA targeting S1pr1 (Dharmacon, # D-001810) using 2 μl/ml Lipofectamine 2000 (Invitrogen, #11668027) or Lipofectamine RNAiMAX (Invitrogen, #13778075) in OptiMEM (GIBCO) as previously described (Rudini et al., 2008). Cells were cultured in complete medium for a further 48 h (S1pr1 siRNA) or 78 h (Cdh5 siRNA) before starvation (18 h for MS1, 6 h for HUVECs), and subsequent stimulations with VEGFR2 and S1PR1 agonists (hVEGF165, SEW2871, S1P from Cayman Chemicals) and processing for staining. siCONTROL Non-Targeting siRNA Pool (Dharmacon, #D-001810) at the same concentration was used as a control for all the siRNA interference assay.

Protocol tips

HUVECs and MS-1 cells were incubated with 40 nM ON-TARGETplus SMARTpool duplex siRNA targeting Cdh5 (Dharmacon, #L003641; L-041968) or 20 nM siRNA targeting S1pr1 (Dharmacon, # D-001810) using 2 μl/ml Lipofectamine 2000 (Invitrogen, #11668027) or Lipofectamine RNAiMAX (Invitrogen, #13778075) in OptiMEM (GIBCO) as previously described (Rudini et al., 2008). Cells were cultured in complete medium for a further 48 h (S1pr1 siRNA) or 78 h (Cdh5 siRNA) before starvation (18 h for MS1, 6 h for HUVECs), and subsequent stimulations with VEGFR2 and S1PR1 agonists (hVEGF165, SEW2871, S1P from Cayman Chemicals) and processing for staining. siCONTROL Non-Targeting siRNA Pool (Dharmacon, #D-001810) at the same concentration was used as a control for all the siRNA interference assay.

Publication protocol

HUVECs and MS-1 cells were incubated with 40 nM ON-TARGETplus SMARTpool duplex siRNA targeting Cdh5 (Dharmacon, #L003641; L-041968) or 20 nM siRNA targeting S1pr1 (Dharmacon, # D-001810) using 2 μl/ml Lipofectamine 2000 (Invitrogen, #11668027) or Lipofectamine RNAiMAX (Invitrogen, #13778075) in OptiMEM (GIBCO) as previously described (Rudini et al., 2008). Cells were cultured in complete medium for a further 48 h (S1pr1 siRNA) or 78 h (Cdh5 siRNA) before starvation (18 h for MS1, 6 h for HUVECs), and subsequent stimulations with VEGFR2 and S1PR1 agonists (hVEGF165, SEW2871, S1P from Cayman Chemicals) and processing for staining. siCONTROL Non-Targeting siRNA Pool (Dharmacon, #D-001810) at the same concentration was used as a control for all the siRNA interference assay.

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