Invitrogen

PRAS40 Monoclonal Antibody (73P21)

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PRAS40 Antibody in Immunocytochemistry (ICC/IF) — Immunofluorescent analysis of PRAS40 was done on 70% confluent log phase HeLa cells. The cells were fixed with 4% paraformaldehyde for 15 minutes, permeabilized with 0.25% Triton™ X-100 for 10 minutes, and blocked with 5% BSA for 1 hour at room temperature. The cells were labeled with PRAS40 Mouse monoclonal Antibody (Product # AHO1031) at 2 µg/mL in 1% BSA and incubated for 3 hours at room temperature and then labeled with Alexa Fluor 488 Rabbit Anti-Mouse IgG Secondary Antibody (Product # A-11059) at a dilution of 1:400 for 30 minutes at room temperature (Panel a: green). Nuclei (Panel b: blue) were stained with SlowFade® Gold Antifade Mountant DAPI (Product # S36938). F-actin (Panel c: red) was stained with Alexa Fluor 594 Phalloidin (Product # A12381). Panel d is a merged image showing cytoplasmic localization. Panel e shows no primary antibody control. The images were captured at 20X magnification.

PRAS40 Antibody in Immunohistochemistry (Paraffin) (IHC (P)) — Immunohistochemistry analysis of PRAS40 showing staining in the cytoplasm and nucleus of paraffin-embedded human heart tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a PRAS40 monoclonal antibody (Product # AHO1031) diluted in 3% BSA-PBS at a dilution of 1:20 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

PRAS40 Antibody in Western Blot (WB) — Extract prepared from human HeLa cells was resolved by SDS-PAGE on a 4-20% polyacrylamide gel and transferred to PVDF. The membrane was blocked with a 5% milk-TBST buffer and then incubated with this PRAS40 monoclonal antibody (clone 73P21) at a concentration of 0.50 µg/mL in a 5% milk-TBST buffer for two hours at room temperature. After washing, the membrane was incubated with goat F (ab')2 anti-mouse IgG alkaline phosphatase (Product # AMI4405) and the signal was detected using the Tropix WesternStar™method. “

PRAS40 Antibody — Figure 4 Co-treatment of Hsp90 and FAK inhibitors reduces canonical survival signaling and induces apoptotic markers in H460 cells A) H460 cells treated for 24h with Hsp90 and FAK inhibitors were used for western blot analysis of FAK, Akt and Erk1/2 signaling. B-D) Quantifications of western blots from A. The phosphorylated proteins, FAK pY 397 (B), Akt pS 473 (C), Erk1/2 pT 202 /Y 204 (D), were normalized to Hsp90, total Akt and Erk1/2, respectively. Analyses were conducted by one-way ANOVA and Dunnett's multiple comparison post-test to compare all columns to control (* p<.05). E) Western blots were conducted for apoptotic markers after 24h of co-treatment cells with FAK and Hsp90 inhibitors alone, or in combination. Western blots are representative of 3 different experiments. Western blot densitometry quantifications are the averaged values from 3 different experiments. {TM}

Product Details

AHO1031

Applications	Tested Dilution	Publications
Western Blot (WB)	Assay-dependent	View 6 publications 6 publications
Immunohistochemistry (Paraffin) (IHC (P))	1:10-1:100	-
Immunocytochemistry (ICC/IF)	1:100-1:500	View 1 publication 1 publication
Immunoprecipitation (IP)	Assay-dependent	View 1 publication 1 publication

Product Specifications
Species Reactivity	Human
Published species	Human, Mouse, Rat
Host/Isotype	Mouse / IgG1, kappa
Class	Monoclonal
Type	Antibody
Clone	73P21
Immunogen	Recombinant full length human PRAS40 expressed in E. coli. View immunogen
Conjugate	Unconjugated
Form	Liquid
Concentration	0.5 mg/mL
Purification	purified
Storage buffer	PBS with 1% BSA
Contains	0.1% sodium azide
Storage conditions	-20°C
RRID	AB_2536321

Target Information

The Akt signaling pathway contributes to the regulation of apoptosis after a variety of cell death signals. AKT1S1, also known as PRAS40, is a proline-rich substrate of the kinase AKT1 and is thought to play a role in neuroprotection mediated by nerve growth factor (NGF) after transient focal cerebral ischemia. AKT1S1 is also a substrate and potential regulator of mammalian target of rapamycin (mTOR), a serine/threonine kinase that regulates cell growth and cell cycle, and a negative regulator of autophagy. Treatment with the insulin-like growth factor-1 (IGF1) can induce the phosphorylation of AKT1S1 via the PI3K/AKT signaling pathway in PC12 cells.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

Product References

Location-specific inhibition of Akt reveals regulation of mTORC1 activity in the nucleus.

Nature communications

Zhou X,Zhong Y,Molinar-Inglis O,Kunkel MT,Chen M,Sun T,Zhang J,Shyy JY,Trejo J,Newton AC,Zhang J

AHO1031 was used in Immunocytochemistry-immunoflourescence to reveal a mode of regulation of mTORC1 that is distinct from its lysosomal activation, which controls mTORC1 activity in the nuclear compartment.

Mon Nov 30 00:00:00 EST 2020

Combination of heat shock protein 90 and focal adhesion kinase inhibitors synergistically inhibits the growth of non-small cell lung cancer cells.

Oncoscience

Webber PJ,Park C,Qui M,Ramalingam SS,Khuri FR,Fu H,Du Y

Published figure using PRAS40 monoclonal antibody (Product # AHO1031) in Western Blot

Tue Oct 27 00:00:00 EDT 2015

Characterization of the activity of the PI3K/mTOR inhibitor XL765 (SAR245409) in tumor models with diverse genetic alterations affecting the PI3K pathway.

Molecular cancer therapeutics

Yu P,Laird AD,Du X,Wu J,Won KA,Yamaguchi K,Hsu PP,Qian F,Jaeger CT,Zhang W,Buhr CA,Shen P,Abulafia W,Chen J,Young J,Plonowski A,Yakes FM,Chu F,Lee M,Bentzien F,Lam ST,Dale S,Matthews DJ,Lamb P,Foster P

AHO1031 was used in western blot to study the anti-tumor effects of a pan class I PI3 kinase/mTOR inhibitor in cells with a genetically diverse range of PI3 kinase pathway modifications.

Thu May 01 00:00:00 EDT 2014

Effects of adding exercise to a 16-week very low-calorie diet in obese, insulin-dependent type 2 diabetes mellitus patients.

The Journal of clinical endocrinology and metabolism

Snel M,Gastaldelli A,Ouwens DM,Hesselink MK,Schaart G,Buzzigoli E,Frölich M,Romijn JA,Pijl H,Meinders AE,Jazet IM

Sun Jul 01 00:00:00 EDT 2012

Phosphorylation of PRAS40 on Thr246 by PKB/AKT facilitates efficient phosphorylation of Ser183 by mTORC1.

Cellular signalling

Nascimento EB,Snel M,Guigas B,van der Zon GC,Kriek J,Maassen JA,Jazet IM,Diamant M,Ouwens DM

Tue Jun 01 00:00:00 EDT 2010

Phosphorylation of PRAS40 on Thr246 by PKB/AKT facilitates efficient phosphorylation of Ser183 by mTORC1.

Cellular signalling

Nascimento EB,Snel M,Guigas B,van der Zon GC,Kriek J,Maassen JA,Jazet IM,Diamant M,Ouwens DM

Tue Jun 01 00:00:00 EDT 2010

The proline-rich Akt substrate of 40 kDa (PRAS40) is a physiological substrate of mammalian target of rapamycin complex 1.

The Journal of biological chemistry

Oshiro N,Takahashi R,Yoshino K,Tanimura K,Nakashima A,Eguchi S,Miyamoto T,Hara K,Takehana K,Avruch J,Kikkawa U,Yonezawa K

AHO1031 was used in western blot to show that proline-rich Akt substrate of 40 kilodaltons is a raptor-binding protein phosphorylated directly by mammalian target of rapamycin complex 1

Fri Jul 13 00:00:00 EDT 2007

Insulin signalling to mTOR mediated by the Akt/PKB substrate PRAS40.

Nature cell biology

Vander Haar E,Lee SI,Bandhakavi S,Griffin TJ,Kim DH

Thu Mar 01 00:00:00 EST 2007

Insulin signalling to mTOR mediated by the Akt/PKB substrate PRAS40.

Nature cell biology

Vander Haar E,Lee SI,Bandhakavi S,Griffin TJ,Kim DH

Thu Mar 01 00:00:00 EST 2007

Insulin signalling to mTOR mediated by the Akt/PKB substrate PRAS40.

Nature cell biology

Vander Haar E,Lee SI,Bandhakavi S,Griffin TJ,Kim DH

Thu Mar 01 00:00:00 EST 2007

Insulin signalling to mTOR mediated by the Akt/PKB substrate PRAS40.

Nature cell biology

Vander Haar E,Lee SI,Bandhakavi S,Griffin TJ,Kim DH

Thu Mar 01 00:00:00 EST 2007

Bioinformatics

Protein Aliases: 40 kDa proline-rich AKT substrate; AKT1 substrate 1 (proline rich); AKT1 substrate 1 (proline-rich); AKT1S1; OTTHUMP00000196756; OTTHUMP00000196760; Proline-rich AKT1 substrate 1

Gene Aliases: AKT1S1; Lobe; PRAS40

UniProt ID: (Human) J3KPM3

Entrez Gene ID: (Human) 84335

Function(s)

negative regulation of protein kinase activity negative regulation of TOR signaling regulation of neuron apoptotic process negative regulation of cell size neurotrophin TRK receptor signaling pathway regulation of cellular response to heat

Process(es)

protein binding

PRAS40 Monoclonal Antibody (73P21)

Product Details

AHO1031

Western Blot (WB)

Immunohistochemistry (Paraffin) (IHC (P))

Immunocytochemistry (ICC/IF)

Immunoprecipitation (IP)

Species Reactivity

Published species

Host/Isotype

Class

Type

Clone

Immunogen

Conjugate

Form

Concentration

Purification

Storage buffer

Contains

Storage conditions

RRID

Target Information

Bioinformatics

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