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Phospho-AKT (Ser473) Monoclonal antibody

Phospho-AKT (Ser473) Monoclonal Antibody for WB, IHC, FC (Intra), ELISA
Cat No. 66444-1-Ig

产品说明书

CloneNo. 1C10B8
引用文献(1267)

宿主/亚型

Mouse / IgG1

种属反应性

human, mouse, rat and More (7)

应用

WB, IHC, FC (Intra), ELISA and More (3)

AKT (Ser473), P AKT, P AKT (Ser473), P-AKT, P-AKT (Ser473)

缓冲液配方:  PBS and Azide
PBS and Azide
PBS Only
规格价格库存


经过测试的应用

Positive WB detected inCalyculin A treated PC-3 cells, Calyculin A treated NIH/3T3 cells, Calyculin A treated HEK-293T cells, HSC-T6 cells, TPA treated Jurkat cells, Calyculin A treated HSC-T6 cells
Positive IHC detected inhuman breast cancer tissue, Calyculin A treated Jurkat cells, human colon cancer tissue
Note: suggested antigen retrieval with TE buffer pH 9.0; (*) Alternatively, antigen retrieval may be performed with citrate buffer pH 6.0
Positive FC (Intra) detected inCalyculin A treated PC-3 cells
Planning an IHC experiment? We recommend our IHCeasy AKT Ready-To-Use IHC Kit. AKT primary antibody included.

推荐稀释比

应用推荐稀释比
Western Blot (WB)WB : 1:2000-1:10000
Immunohistochemistry (IHC)IHC : 1:100-1:400
Flow Cytometry (FC) (INTRA)FC (INTRA) : 0.50 ug per 10^6 cells in a 100 µl suspension
It is recommended that this reagent should be titrated in each testing system to obtain optimal results.
Sample-dependent, Check data in validation data gallery.

产品信息

66444-1-Ig targets Phospho-AKT (Ser473) in WB, IHC, IF, FC (Intra), IP, ELISA, IHC-IF applications and shows reactivity with human, mouse, rat samples.

经测试应用 WB, IHC, FC (Intra), ELISA Application Description
文献引用应用WB, IHC, IF, IP, IHC-IF
经测试反应性 human, mouse, rat
文献引用反应性human, mouse, rat, pig, rabbit, monkey, chicken, zebrafish, sheep, duck
免疫原 Peptide 种属同源性预测
宿主/亚型 Mouse / IgG1
抗体类别 Monoclonal
产品类型 Antibody
全称 v-akt murine thymoma viral oncogene homolog 1
别名 AKT (Ser473), P AKT, P AKT (Ser473), P-AKT, P-AKT (Ser473)
观测分子量 60-62 kDa
GenBank蛋白编号NM_005163
基因名称 AKT1
Gene ID (NCBI) 207
RRIDAB_2782958
偶联类型 Unconjugated
形式 Liquid
纯化方式Protein A purification
UNIPROT IDP31749
储存缓冲液 PBS with 0.02% sodium azide and 50% glycerol, pH 7.3.
储存条件Store at -20°C. Stable for one year after shipment. Aliquoting is unnecessary for -20oC storage.

背景介绍

1) What is AKT?

The serine/threonine kinase B AKT pathway (also known as the PI3K-Akt pathway) plays a vital role in the regulation of cellular processes, including cell proliferation, survival, and growth - processes that are essential for oncogenesis. Mutation of the regulator proteins PI3K and PTEN causes uncontrolled disruption within the PI3-kinase pathway, leading to the development of human cancers (1,2; see also AKT pathway poster for more details).

2) phospho-AKT and FAQs

A)  What is the best way to normalize phosphorylated proteins analyzed by western blot?
Normalize phospho-AKT and total AKT with your loading control (e.g. Actin, tubulin), then calculate the phospho/total ratio using these normalized values.  
Put more simply:
1. Calculate the ratio of band intensities of a phospho-AKT band: the loading control.
2. Calculate the ratio of band intensities of total AKT: loading control.
3. Divide ratio obtained #1 by #2 to obtain a normalized value for comparison among different conditions. This procedure allows one to distinguish between a change in AKT expression and a change in the ratio of phospho-AKT.
* If you are looking at the differences in a phospho-AKT expression resulting from an experimental condition (e.g., knockdown), you should also show the expression of total AKT to distinguish between a change in AKT expression (transcription/translation level) and a change in the AKT phosphorylation status.
B) What is the observed molecular weight for AKT and phospho-AKT?
Molecular Weight AKT - 56 kDa
Molecular Weight phospho-AKT - 60 kDa (Figure 1)
  

Figure 1. WB: HEK-293 cell lysate was subjected to SDS PAGE followed by western blot with 60203-2-Ig (AKT antibody) and 66444-1-Ig (AKT-phospho-S473 antibody) at a dilution of 1:4000 incubated at room temperature for 1.5 hours.

C) Are there any special WB conditions to optimize staining of a phospho-AKT?
Since this is a phosphorylated protein, 5% BSA is recommended over non-fat milk as a blocking agent.
D) What are good positive and negative controls for a phospho-AKT?
- Positive Control: HEK293 cells
- Negative Control: Treatment with PI3K inhibitors (e.g. wortmannin)
E) What species does this antibody react with?

Our internal testing has confirmed that it reacts with the human and mouse forms of phospho-AKT.Reactivity with the human form is also supported by the literature's citations of this antibody.

References:

1. Perturbations of the AKT signaling pathway in human cancer.
2. Targeting the PI3K-Akt pathway in human cancer: rationale and promise.

实验方案

Product Specific Protocols
WB protocol for Phospho-AKT (Ser473) antibody 66444-1-IgDownload protocol
IHC protocol for Phospho-AKT (Ser473) antibody 66444-1-IgDownload protocol
Standard Protocols
Click here to view our Standard Protocols

发表文章

SpeciesApplicationTitle
humanWB

Signal Transduct Target Ther

Circulating tumor cells shielded with extracellular vesicle-derived CD45 evade T cell attack to enable metastasis

Authors - Chuan Yang

Adv Mater

Targeted Macrophage CRISPR-Cas13 Mrna Editing in Immunotherapy for Tendon Injury

Authors - Shuo Wang
mouseWB

Cell Metab

Disrupted methionine cycle triggers muscle atrophy in cancer cachexia through epigenetic regulation of REDD1

Authors - Kai Lin
mouseWB,IHC

Cell Res

In vivo self-assembled small RNAs as a new generation of RNAi therapeutics.

Authors - Zheng Fu
mouseIF

Cell Res

Inhibiting Hv1 channel in peripheral sensory neurons attenuates chronic inflammatory pain and opioid side effects.

Authors - Qiansen Zhang
human,mouseWB

Nat Commun

Genome-wide enhancer-gene regulatory maps link causal variants to target genes underlying human cancer risk

Authors - Pingting Ying
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