AKT 1/2/3 antibody (AA 401-479)

Catalog No. ABIN725195

Product Details anti-AKT 1/2/3 Antibody

Target: AKT 1/2/3

Binding Specificity: AA 401-479

Reactivity: Human, Mouse, Rat, Cow, Pig

Host: Rabbit

Clonality: Polyclonal

Conjugate: This AKT 1/2/3 antibody is un-conjugated

Application: Western Blotting (WB), Immunofluorescence (Cultured Cells) (IF (cc)), Immunofluorescence (Paraffin-embedded Sections) (IF (p)), Flow Cytometry (FACS), ELISA, Immunohistochemistry (Paraffin-embedded Sections) (IHC (p)), Immunohistochemistry (Frozen Sections) (IHC (fro))

Cross-Reactivity: Cow, Human, Mouse, Pig, Rat

Predicted Reactivity: Dog,Cow,Sheep,Chicken

Purification: Purified by Protein A.

Immunogen: KLH conjugated synthetic peptide derived from human AKT1

Isotype: IgG

Application Details

Application Notes: WB 1:300-5000
ELISA 1:500-1000
FCM 1:20-100
IHC-P 1:200-400
IHC-F 1:100-500
IF(IHC-P) 1:50-200
IF(IHC-F) 1:50-200
IF(ICC) 1:50-200

Restrictions: For Research Use only

Independent Validation (FACS)

Validation #029769 (Flow Cytometry)

by: Flow Cytometry & Cell Separation Facility, Purdue University

No.: #029769

Date: 07/22/2014

Lot Number: 131113

Method validated: Flow Cytometry

Positive Control: MDA-MB-468 cells - lower expression

Notes: A strong specific signal is observed in both of the positive control cells stained with anti-AKT1 plus secondary antibody compared with isotype, secondary only and unstained cells.

Validation Images

Figure 1: Positive control (high expression) MCF7 cells. The red histogram is unstained cells, the blue histogram is cells stained with secondary antibody alone, the green histogram is cells stained with rabbit IgG isotype control antibody plus secondary antibody and the orange histogram is cells stained with anti-AKT1 plus secondary antibody. The secondary antibody is a goat anti-rabbit IgG-FITC (Jackson Immunoresearch).

Figure 2: Positive control (lower expression) MDA-MB-468 cells. The red histogram is unstained cells, the blue histogram is cells stained with secondary antibody alone, the green histogram is cells stained with rabbit IgG isotype control antibody plus secondary antibody and the orange histogram is cells stained with anti-AKT1 plus secondary antibody. The secondary antibody is a goat anti-rabbit IgG-FITC (Jackson Immunoresearch).

Full Methods

Primary Antibody:

• Antigen: V-Akt Murine Thymoma Viral Oncogene Homolog 1 (AKT1)
• Catalog number: ABIN725195
• Lot number: 131113
• Dilution: 1 µg in 100 µL 1X PBS containing 0.5% BSA

Secondary Antibody:

• Antibody: Goat anti-rabbit IgG-FITC
• Lot number: 115629
• Dilution: 1:200 in 1X PBS containing 0.5% BSA

Full Protocol:

• Control cells were cultured in DMEM + 10% FBS. - Control cells were washed once with phosphate-buffered saline (PBS) and harvested with a non-enzymatic cell dissociation solution (Cellstripper, Mediatech, Inc).
• Detached cells were washed and resuspended in 0.5 mL 1X PBS containing 0.5% BSA.
• An equivalent amount of pre-warmed 4% paraformaldehyde was added and the cells were incubated for 10 min at 37 °C.
• Cells were washed in 1X PBS containing 0.5% BSA and resuspended in 1 mL of ice-cold 90% methanol for 30 min on ice.
• Cells were spun out of the methanol and washed twice by resuspending in 1X PBS containing 0.5% BSA.
• Cells were resuspended in 100 µL 1X PBS containing 0.5% BSA: - unstained cells - secondary antibody alone - isotype control antibody + secondary antibody - primary antibody + secondary antibody
• Cells were incubated for 30 min on ice.
• Labeled cells were washed twice in PBS containing 0.5% BSA.
• Cells were resuspended with 1X PBS containing 0.5% BSA + 10% goat serum and incubated for 15 min at room temperature.
• Goat anti-rabbit IgG-FITC secondary antibody (Jackson Immunoresearch) was added at a 1:200 dilution. The cells were incubated for 30 min in the dark on ice.
• Labeled cells were washed twice in PBS containing 0.5% BSA.
• Cells were analyzed on a FACSAria III (BD Biosciences) using a blue laser (488 nm excitation / 525 nm emission).

Experimental Notes: Nothing to note.

Handling

Format: Liquid

Concentration: 1 μg/μL

Buffer: 0.01M TBS( pH 7.4) with 1 % BSA, 0.02 % Proclin300 and 50 % Glycerol.

Preservative: ProClin

Precaution of Use: This product contains ProClin: a POISONOUS AND HAZARDOUS SUBSTANCE, which should be handled by trained staff only.

Storage: 4 °C,-20 °C

Storage Comment: Shipped at 4°C. Store at -20°C for one year. Avoid repeated freeze/thaw cycles.

Expiry Date: 12 months

References for anti-AKT 1/2/3 antibody (ABIN725195)

Liu, Liu, Liu, Ma, Zhao: "Surgical incision induces learning impairment in mice partially through inhibition of the brain-derived neurotrophic factor signaling pathway in the hippocampus and amygdala." in: Molecular pain, Vol. 14, pp. 1744806918805902, (2019) (PubMed).

Zhang, Zhao, Zhang, Hao, Yu, Min, Li, Ma, Chen, Yi, Tang, Meng, Liu, Wang, Shen, Zhang: "Decrease in male mouse fertility by hydrogen sulfide and/or ammonia can Be inheritable." in: Chemosphere, Vol. 194, pp. 147-157, (2018) (PubMed).

Zhao, Zhang, Liu, Zhang, Hao, Li, Chen, Shen, Tang, Min, Meng, Wang, Yi, Zhang: "Hydrogen Sulfide and/or Ammonia Reduces Spermatozoa Motility through AMPK/AKT Related Pathways." in: Scientific reports, Vol. 6, pp. 37884, (2016) (PubMed).

Zhang, Huang, Guo, Gou, Hinsdale, Lloyd, Liu: "MicroRNA-26b Modulates the NF-κB Pathway in Alveolar Macrophages by Regulating PTEN." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 195, Issue 11, pp. 5404-14, (2015) (PubMed).

Liang, Zeng, Wang, Shen, Li, Ma, Ci, Yu, Jia, Sun, Liu, Liu, Li, Yu, Chen, Jia: "Histone demethylase RBP2 induced by Helicobactor Pylori CagA participates in the malignant transformation of gastric epithelial cells." in: Oncotarget, Vol. 5, Issue 14, pp. 5798-807, (2015) (PubMed).

Li, Liu, Wang, Sun, Ding, Sun, Han, Wang: "Follistatin could promote the proliferation of duck primary myoblasts by activating PI3K/Akt/mTOR signalling." in: Bioscience reports, Vol. 34, Issue 5, (2014) (PubMed).

Xu, Huang, Xie, Lin, Guo, Zhuang, Lin, Zhou, Li, Huang, Wei, Man, Zhang: "The expression of cytoglobin as a prognostic factor in gliomas: a retrospective analysis of 88 patients." in: BMC cancer, Vol. 13, pp. 247, (2013) (PubMed).

Tang, Liu, Dong, Li, Li, Lin, Zheng, Hou, Li: "The cardioprotective effects of citric Acid and L-malic Acid on myocardial ischemia/reperfusion injury." in: Evidence-based complementary and alternative medicine : eCAM, Vol. 2013, pp. 820695, (2013) (PubMed).

Target Details for AKT 1/2/3

Target: AKT 1/2/3

Alternative Name: AKT1/2/3 (AKT 1/2/3 Products)

Background:

Synonyms: AKT, PKB, RAC, CWS6, PRKBA, PKB-ALPHA, RAC-ALPHA, RAC-alpha serine/threonine-protein kinase, Protein kinase B, Protein kinase B alpha, PKB alpha, Proto-oncogene c-Akt, RAC-PK-alpha, AKT1, RAC-gamma serine/threonine-protein kinase, Protein kinase Akt-3, Protein kinase B gamma, PKB gamma, RAC-PK-gamma, STK-2, PKBG

Background: AKT1-specific substrates have been recently identified, including palladin (PALLD), which phosphorylation modulates cytoskeletal organization and cell motility, prohibitin (PHB), playing an important role in cell metabolism and proliferation, and CDKN1A, for which phosphorylation at 'Thr-145' induces its release from CDK2 and cytoplasmic relocalization. These recent findings indicate that the AKT1 isoform has a more specific role in cell motility and proliferation. Phosphorylates CLK2 thereby controlling cell survival to ionizing radiation.

Gene ID: 207

UniProt: P31749