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AB253299

AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit

4

(1 Review)

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(1 Publication)

AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit is a single-wash 90-min Simplestep used to quantify AKT 1/2/3 pS473 + AKT 1/2/3 Total with a sensitivity of 1 µg/mL. The assay uses a simple mix-wash-read protocol with just one incubation and one wash step.

- Colorimetric Sandwich ELISA - 450 nm readout : works on any standard plate reader
4 Images
Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)
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Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)

Example of a typical AKT 1/2/3 (PS473) and AKT1 Total recombinant protein standard curve. The proportion of total protein that is phosphorylated is unknown - data is indicative only. Raw data values are shown in the table. Background-subtracted data values (mean +/- SD) are graphed.

Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)
  • sELISA

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Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)

Typical cell lysate dilution series.

Example of AKT 1/2/3 (pS473) and AKT 1/2/3 Total control lysate dilution series in 1X Cell Extraction Buffer PTR.

Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)
  • sELISA

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Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)

Induction of AKT 1/2/3 (pS473) phosphorylation in MCF-7 cells in response to insulin treatment, and levels of AKT 1/2/3 Total in the same samples.

MCF-7 cells were cultured in 96-well tissue culture plates, serum-starved (2 hr) and treated (30 min) with a dose-range of insulin before cell lysis. Data from duplicate measurements of AKT 1/2/3 (pS473) are plotted and compared against AKT 1/2/3 Total protein levels.

Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)
  • sELISA

Unknown

Sandwich ELISA - AKT 1/2/3 pS473 + AKT 1/2/3 Total ELISA Kit (AB253299)

Cell line analysis for AKT 1/2/3 Total from preparations of 17,500 cells/well cell extracts.

Data from triplicate measurements (mean +/- SD) are plotted.

Key facts

Sample types

Cell Lysate

Reacts with

Mouse, Human

Results type

Semi-Quantitative

Sensitivity

< 1 µg/mL

Range

0.05 - 3.12 %

Assay Platform

Pre-coated microplate (12 x 8 well strips)

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Primary Antibodies

AB38449

Anti-AKT (phospho T308) antibody

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-AKT (phospho T308) antibody (AB38449)

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Secondary Antibodies

AB150077

Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488)

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Primary Antibodies

AB9485

Anti-GAPDH antibody - Loading Control

BOND RX™ Validated|Lab Essentials

Immunocytochemistry/ Immunofluorescence - Anti-GAPDH antibody - Loading Control (AB9485)

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Reactivity data

{ "title": "Reactivity Data", "filters": { "stats": ["", "Reactivity", "Dilution Info", "Notes"] }, "values": { "ELISA": { "reactivity":"TESTED_AND_REACTS", "dilution-info":"", "notes":"<p></p>" } } }
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Product details

AKT 1/2/3 pS473 + AKT 1/2/3 Total (ab253299) is a single-wash 90 min sandwich ELISA designed for the semi-quantitative measurement of total AKT 1/2/3 pS473 + AKT 1/2/3 total protein in human and mouse cell lysates. It uses our proprietary SimpleStep ELISA® technology. Semi-quantitate AKT 1/2/3 pS473 + AKT 1/2/3 total with 1 μg/mL sensitivity.

SimpleStep ELISA® technology employs capture antibodies conjugated to an affinity tag that is recognized by the monoclonal antibody used to coat our SimpleStep ELISA® plates. This approach to sandwich ELISA allows the formation of the antibody-analyte sandwich complex in a single step, significantly reducing assay time. See the SimpleStep ELISA® protocol summary in the image section for further details. Our SimpleStep ELISA® technology provides several benefits:

-Single-wash protocol reduces assay time to 90 minutes or less
-High sensitivity, specificity and reproducibility from superior antibodies
-Fully validated in biological samples
-96-wells plate breakable into 12 x 8 wells strips

A 384-well SimpleStep ELISA® microplate (ab203359) is available to use as an alternative to the 96-well microplate provided with SimpeStep ELISA® kits.

AKT isoforms belong to a conserved family of serine/threonine protein kinases. AKT plays a central role in cell survival and inhibiting apoptosis, by phosphorylating and inactivating several targets. AKT is activated by insulin and various other agonists, which promote transport to the plasma membrane, where the activation loop of AKT is phosphorylated at Thr308 by PDK1. The carboxy terminus is phosphorylated at Ser473 by mammalian target of rapamycin complex 2 (TORC2).

AKT substrates include Bad, forkhead transcription factors, and caspase-9. Another vital AKT function is the regulation of glycogen synthesis through phosphorylation of GSK-3. AKT phosphorylation also prevents GSK-3β mediated phosphorylation and degradation of cyclin D1, and negatively regulates the cyclin dependent kinase inhibitors p27 Kip and p21 Waf1/CIP1. AKT also plays a critical role in the regulation of TORC1 and tuberin (TSC2).

REACH authorisation
Abcam has not and does not intend to apply for the REACH Authorisation of customers' uses of products that contain European Authorisation list (Annex XIV) substances.
It is the responsibility of our customers to check the necessity of application of REACH Authorisation, and any other relevant authorisations, for their intended uses.

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Precision

[ { "reproducibilityType": "Inter", "sample": "AKT (pS473)", "replicates": 3, "mean": null, "standardDeviation": null, "coefficientOfVariability": "5.3" }, { "reproducibilityType": "Inter", "sample": "AKT Total", "replicates": 3, "mean": null, "standardDeviation": null, "coefficientOfVariability": "9.5" }, { "reproducibilityType": "Intra", "sample": "AKT (pS473)", "replicates": 6, "mean": null, "standardDeviation": null, "coefficientOfVariability": "8.7" }, { "reproducibilityType": "Intra", "sample": "AKT Total", "replicates": 6, "mean": null, "standardDeviation": null, "coefficientOfVariability": "1.8" } ]
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What's included?

{ "values": { "1x96Tests": { "sellingSize": "1 x 96 Tests", "publicAssetCode":"ab253299-1x96Tests", "assetComponentDetails": [ { "size":"1 x 1.5 mL", "name":"AKT 1/2/3 Total Capture Antibody", "number":"AB253299-CMP03", "productcode":"" }, { "size":"1 x 1.5 mL", "name":"AKT 1/2/3 (pS473) Capture Antibody", "number":"AB253299-CMP05", "productcode":"" }, { "size":"1 x 15 mL", "name":"10X Wash Buffer PT", "number":"AB253299-CMP02", "productcode":"" }, { "size":"1 x 1 Vial", "name":"Lyophilized AKT Control Lysate", "number":"AB253299-CMP08", "productcode":"" }, { "size":"1 x 1 mL", "name":"50X Cell Extraction Enhancer Solution", "number":"AB253299-CMP07", "productcode":"" }, { "size":"1 x 1 Unit", "name":"Plate Seal", "number":"AB253299-CMP09", "productcode":"" }, { "size":"1 x 12 mL", "name":"TMB Substrate", "number":"AB253299-CMP11", "productcode":"" }, { "size":"1 x 1 Unit", "name":"SimpleStep Pre-Coated 96-Well Microplate (ab206978)", "number":"AB253299-CMP01", "productcode":"" }, { "size":"1 x 1.5 mL", "name":"AKT 1/2/3 Total Detector Antibody", "number":"AB253299-CMP04", "productcode":"" }, { "size":"1 x 1.5 mL", "name":"AKT 1/2/3 (pS473) Detector Antibody", "number":"AB253299-CMP06", "productcode":"" }, { "size":"1 x 12 mL", "name":"Stop Solution", "number":"AB253299-CMP10", "productcode":"" }, { "size":"1 x 12 mL", "name":"5X Cell Extraction Buffer PTR", "number":"AB253299-CMP12", "productcode":"" } ] } } }
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Properties and storage information

Shipped at conditions
Blue Ice
Appropriate short-term storage conditions
+4°C
Appropriate long-term storage conditions
+4°C
Storage information
+4°C
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Supplementary information

This supplementary information is collated from multiple sources and compiled automatically.

AKT 1/2/3 also known as protein kinase B (PKB) consists of three isoforms; AKT1 AKT2 and AKT3. These proteins are serine/threonine-specific kinases with a molecular weight around 56 kDa. They are phosphorylated at Ser473 which regulates their activity. AKT proteins are expressed ubiquitously but the expression levels can vary across different tissues. AKT1 is primarily found in most tissues AKT2 is mostly present in insulin-responsive tissues like liver muscle and adipose tissue while AKT3 is largely detected in the brain.
Biological function summary

AKT proteins perform critical roles in key signaling pathways such as the PI3K/AKT pathway which is important for cellular processes like growth proliferation and survival. AKT activation requires phosphorylation at two sites: Thr308 in the activation loop and Ser473 in the hydrophobic motif. These kinases attend a complex signaling network involving direct phosphorylation of multiple downstream targets influencing numerous cellular processes. AKT1 AKT2 and AKT3 might have overlapping as well as unique functions in vivo reflecting their different tissue distributions and substrate specificities.

Pathways

AKT proteins are central components of the insulin signaling pathway and the PI3K/AKT/mTOR pathway. These pathways play essential roles in regulating metabolism cell survival and growth. AKT interacts with various other proteins such as mTOR and FOXO transcription factors. In the PI3K/AKT/mTOR pathway AKT acts downstream of PI3K to activate or inhibit targets that regulate protein synthesis metabolism and apoptosis.

AKT dysregulation is associated with many cancers and metabolic diseases like diabetes. Aberrant activation of the PI3K/AKT/mTOR pathway contributes to oncogenesis as it promotes cell survival and proliferation. In metabolic diseases altered AKT activity affects insulin signaling leading to insulin resistance and type 2 diabetes. AKT is also linked with proteins such as PTEN which serve as tumor suppressors by inhibiting the PI3K/AKT pathway showing the importance of balancing AKT activity for normal cellular function.
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Product protocols

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Target data

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Publications (1)

Recent publications for all applications. Explore the full list and refine your search

Nature communications 14:2271 PubMed37080971

2023

Glucocorticoid activation of anti-inflammatory macrophages protects against insulin resistance.

Applications

Unspecified application

Species

Unspecified reactive species

Giorgio Caratti,Ulrich Stifel,Bozhena Caratti,Ali J M Jamil,Kyoung-Jin Chung,Michael Kiehntopf,Markus H Gräler,Matthias Blüher,Alexander Rauch,Jan P Tuckermann
View all publications
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Product promise

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