Western blot analysis of extracts from serum-starved HeLa cells, untreated or treated with IGF-1 (50 ng/ml, 30 minutes) and λ phosphatase, using Phospho-Rictor (Thr1135) (D30A3) Rabbit mAb (upper) or Rictor (53A2) Rabbit mAb #2114 (lower).
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Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
Phospho-Rictor (Thr1135) (D30A3) Rabbit mAb detects endogenous levels of rictor protein only when phosphorylated at Thr1135.
Human, Mouse, Monkey
Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to the sequence surrounding Thr1135 of human Rictor protein.
Cell growth is a fundamental biological process whereby cells accumulate mass and increase in size. The mammalian TOR (mTOR) pathway regulates growth by coordinating energy and nutrient signals with growth factor-derived signals (1). mTOR is a large protein kinase with two different complexes. One complex contains mTOR, GβL and raptor, which is a target of rapamycin. The other complex, insensitive to rapamycin, includes mTOR, GβL, Sin1, and rictor (1). The mTOR-rictor complex phosphorylates Ser473 of Akt/PKB in vitro (2). This phosphorylation is essential for full Akt/PKB activation. Furthermore, an siRNA knockdown of rictor inhibits Ser473 phosphorylation in 3T3-L1 adipocytes (3). This complex has also been shown to phosphorylate the rapamycin-resistant mutants of S6K1, another effector of mTOR (4).
Phosphorylation of Thr1135 on rictor was identified at Cell Signaling Technology (CST) using PhosphoScan®, CST's LC-MS/MS platform for phosphorylation site discovery (5). Additional research indicates that rictor is phosphorylated at Thr1135 by p70 S6K, which negatively regulates mTORC2 protein complex as part of a negative feedback mechanism controlling Akt activity (6-8).
Explore pathways + proteins related to this product.
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