Cell Signaling
S6 Ribosomal Protein
2217
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The general aim of this project was to assess protein synthesis in a variety of treatment groups using ribosomal protein s6 (rps6), a downstream component of the MTORC1 pathway. To do this we compared phosphorylated rps6 to total rps6 using western blotting.
Western Blot
Total cell lysates of rat hepatoma cells
1:2000 antibody in 0.1% tween and 1:3 odyssey block: 1X TBS at 4 degrees C overnight.
1:3 odyssey block:1X TBS
1:15,000 Li-Cor IRDye 800CW, 0.1% tween, 0.02% SDS, in 1:3 odyssey block: TBS at room temperature for 1 hour
N/A
Li-Cor Odyssey infrared Imaging system
There was a large distinction between samples that were phosphorylated (indicating active protein synthesis) and samples that were not phosphorylated (indicating halted protein synthesis). These results were consistent with another experiment performed to observe active protein synthesis via phosphorylation
None
Dilution from protocol worked well the first time and two times after. Have a lot of confidence with this antibody
Had no issues in detecting total rps6 in this rat hepatoma cell line.