Microplate assays to measure total and phospho-S6 in whole cells.
Thermo Scientific Pierce S6 In-Cell ELISA Kits are colorimetric and fluorescent microplate assays optimized to measure total and phosphorylated ribosomal S6 protein (phospho-S6) in whole cells.
These ELISA kits for whole cells include a mouse monoclonal anti-S6 antibody, rabbit monoclonal anti-phospho-(Ser235/236) S6 antibody, detection reagents and all buffers required for either colorimetric or near infrared detection. The S6 In-Cell ELISA Kits enable simultaneous detection of both total and phospho-(Ser235/236) S6.
Measures total and phosphorylated ribosomal S6 protein in whole cells using a 96- or 384-well microplates
colorimetric detection kit for use with standard microplate readers
near infrared fluorescent detection kit for use with IR imaging system
Whole-cell detection requires no cell lysis, protein extraction or special sample preparation
Provides better data and higher throughput than quantitative Western blotting
Less costly (one well vs. one lane) than Western blotting
Upon recovery from serum-starvation, phosphorylation of ribosomal S6 protein increases >3-fold in NIH3T3. Relative total and phospho-S6 levels were measured in NIH3T3 cells using the (A) S6 Colorimetric In-Cell ELISA Kit and the (B) S6 Near Infrared In-Cell ELISA Kit. Cells were serum-starved overnight and stimulated with 20% fetal bovine serum 5 minutes immediately prior to fixation.The level of S6 phosphorylation increased 3- to 8-fold (colorimetric detection and near IR detection, respectively) in response to serum treatment where as total S6 protein levels were not significantly affected. Note: Absolute absorbance values cannot be directly compared between phospho and total S6 experiments because they use antibodies with different sensitivities.
Overview of S6 and phosphorylation of S6:
The ribosomal S6 protein is a component of the small subunit of ribsomes (40S) and is involved in the regulation of translation. The activity of S6 is regulated, in part, by the downstream effectors of mTOR, the ribosomal protein S6 kinase (S6K). Phosphorylation of S6 by S6K is important in the regulation of protein synthesis, cell size, cell-cycle progression, glucose homeostasis and survival of newborns. S6K activation and phosphorylation of S6 are implicated in translation efficiency of mRNAs with a 5'-terminal oligopyrimidine tract (TOP mRNAs) in certain physiological conditions. S6 phosphorylation occurs on 5 serine residues (Ser235, Ser236, Ser240, Ser244 and Ser247) within the C-terminus of the protein and are evolutionarily conserved in higher eukaryotes.The S6 In-Cell ELISA Kit enables simultaneous detection of both total and phospho-(Ser235/236) S6.