caveolin-1 siRNA (h)

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	Human bronchial epithelial cells (BEAS-2B; ATCC CRL-9609) were routinely cultured in RPMI base medium supplemented with 10% fetal bovine serum (FBS) and 1× penicillin-streptomycin in 75-cm2 tissue culture flasks at 37°C in the presence of 5% CO2. Cells were grown to approximately 75% confluence and passaged. Cells were discarded after their 9th passage. Cells were plated in a 24-well plate for all assays at a density of 250,000 cells per well. Cells were allowed to grow for approximately 48 h prior to challenge with a medium exchange at 24 h prior to challenge. Serum-starved cells were given RPMI medium lacking FBS 24 h prior to challenge. Transient expression of mCherry fusion proteins occurred through a standardized lipofection-based protocol described by Clark et al. (56). Transient silencing of a specified gene occurred through Lipofectamine LTX with Plus solution via the manufacturer's protocol. Specifically, liposomes were prepared using 1 to 5 μl siRNA (Santa Cruz), 5 μl Plus solution per well, and 5 μl Lipofectamine LTX per well

Protocol tips

Human bronchial epithelial cells (BEAS-2B; ATCC CRL-9609) were routinely cultured in RPMI base medium supplemented with 10% fetal bovine serum (FBS) and 1× penicillin-streptomycin in 75-cm2 tissue culture flasks at 37°C in the presence of 5% CO2. Cells were grown to approximately 75% confluence and passaged. Cells were discarded after their 9th passage. Cells were plated in a 24-well plate for all assays at a density of 250,000 cells per well. Cells were allowed to grow for approximately 48 h prior to challenge with a medium exchange at 24 h prior to challenge. Serum-starved cells were given RPMI medium lacking FBS 24 h prior to challenge. Transient expression of mCherry fusion proteins occurred through a standardized lipofection-based protocol described by Clark et al. (56). Transient silencing of a specified gene occurred through Lipofectamine LTX with Plus solution via the manufacturer's protocol. Specifically, liposomes were prepared using 1 to 5 μl siRNA (Santa Cruz), 5 μl Plus solution per well, and 5 μl Lipofectamine LTX per well

Publication protocol

Human bronchial epithelial cells (BEAS-2B; ATCC CRL-9609) were routinely cultured in RPMI base medium supplemented with 10% fetal bovine serum (FBS) and 1× penicillin-streptomycin in 75-cm2 tissue culture flasks at 37°C in the presence of 5% CO2. Cells were grown to approximately 75% confluence and passaged. Cells were discarded after their 9th passage. Cells were plated in a 24-well plate for all assays at a density of 250,000 cells per well. Cells were allowed to grow for approximately 48 h prior to challenge with a medium exchange at 24 h prior to challenge. Serum-starved cells were given RPMI medium lacking FBS 24 h prior to challenge. Transient expression of mCherry fusion proteins occurred through a standardized lipofection-based protocol described by Clark et al. (56). Transient silencing of a specified gene occurred through Lipofectamine LTX with Plus solution via the manufacturer's protocol. Specifically, liposomes were prepared using 1 to 5 μl siRNA (Santa Cruz), 5 μl Plus solution per well, and 5 μl Lipofectamine LTX per well

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