Phospho-AKT (Ser473) Monoclonal antibody, PBS Only (Capture)

Phospho-AKT (Ser473) Monoclonal Antibody for WB, FC, IHC, Indirect ELISA, Cytometric bead array

Host / Isotype

Mouse / IgG1

Reactivity

human, mouse, rat

Applications

WB, FC, IHC, Indirect ELISA, Cytometric bead array

Conjugate

Unconjugated

CloneNo.

1C10B8

Cat no : 66444-1-PBS

Synonyms

AKT, AKT1, pakt, PKB, PKB ALPHA, PRKBA, Protein kinase B, Proto oncogene c Akt, RAC, RAC ALPHA, RAC PK alpha

---

Validation Data Gallery

• 

  WB analysis using 66444-1-Ig (same clone as 66444-1-PBS)

  Non-treated Jurkat and HEK-293 cells and Calyculin A treated Jurkat and HEK-293 cells were subjected to SDS PAGE followed by western blot with 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:10000 incubated at room temperature for 1.5 hours. This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  WB analysis using 66444-1-Ig (same clone as 66444-1-PBS)

  Non-treated PC-3 and Calyculin A treated PC-3 cells were subjected to SDS PAGE followed by western blot with 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:5000 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with GAPDH antibody as loading control. This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  WB analysis using 66444-1-Ig (same clone as 66444-1-PBS)

  Non-treated HEK-293T and IGF1 or Calyculin A treated HEK-293T cells were subjected to SDS PAGE followed by western blot with 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:5000 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with GAPDH antibody as loading control.. This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  WB analysis using 66444-1-Ig (same clone as 66444-1-PBS)

  Non-treated HSC-T6 and Calyculin A treated HSC-T6 cells were subjected to SDS PAGE followed by western blot with 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:10000 incubated at room temperature for 1.5 hours. This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  WB analysis using 66444-1-Ig (same clone as 66444-1-PBS)

  Non-treated NIH/3T3 and Calyculin A treated NIH/3T3 cells were subjected to SDS PAGE followed by western blot with 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:10000 incubated at room temperature for 1.5 hours. This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  WB analysis using 66444-1-Ig (same clone as 66444-1-PBS)

  Non-treated Jurkat and TPA treated Jurkat cells were subjected to SDS PAGE followed by western blot with 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:5000 incubated at room temperature for 1.5 hours. The membrane was stripped and re-blotted with GAPDH antibody as loading control. This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  IHC staining of human breast cancer using 66444-1-Ig (same clone as 66444-1-PBS)

  Immunohistochemical analysis of paraffin-embedded human breast cancer tissue slide using 66444-1-Ig (AKT-phospho-S473 antibody at dilution of 1:200 (under 10x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0). This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  IHC staining of human breast cancer using 66444-1-Ig (same clone as 66444-1-PBS)

  Immunohistochemical analysis of paraffin-embedded human breast cancer tissue slide using 66444-1-Ig (AKT-phospho-S473 antibody at dilution of 1:200 (under 40x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0). This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  IHC staining of Jurkat using 66444-1-Ig (same clone as 66444-1-PBS)

  Immunohistochemical analysis of paraffin-embedded untreated (left) or Calyculin A treated (right) Jurkat cells slide using 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:8000 (under 10x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0). This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  IHC staining of Jurkat using 66444-1-Ig (same clone as 66444-1-PBS)

  Immunohistochemical analysis of paraffin-embedded untreated (left) or Calyculin A treated (right) Jurkat cells slide using 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:8000 (under 40x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0). This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  IHC staining of human colon cancer using 66444-1-Ig (same clone as 66444-1-PBS)

  Immunohistochemical analysis of paraffin-embedded human colon cancer tissue slide using 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:2000 (under 10x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0). This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  IHC staining of human colon cancer using 66444-1-Ig (same clone as 66444-1-PBS)

  Immunohistochemical analysis of paraffin-embedded human colon cancer tissue slide using 66444-1-Ig (Phospho-AKT (Ser473) antibody) at dilution of 1:2000 (under 40x lens). Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0). This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  FC experiment of PC-3 using 66444-1-Ig (same clone as 66444-1-PBS)

  1X10^6 PC-3 cells untreated (dashed line) or treated with Calyculin A (red) were intracellularly stained with 0.5 ug Anti-Human Phospho-AKT (Ser473) (66444-1-Ig, Clone:1C10B8) and CoraLite®488-Conjugated AffiniPure Goat Anti-Mouse IgG(H+L) at dilution 1:1000, or 0.5 ug Control Antibody (blue). Cells were fixed with 4% PFA and permeabilized with 90% MeOH. This data was developed using the same antibody clone with 66444-1-PBS in a different storage buffer formulation.

• 

  Cytometric bead array in cell lysate using MP50177-1

  Cytometric bead array in cell lysate using MP50177-1, Phospho-AKT(Ser473) Monoclonal Matched Antibody Pair, PBS Only. Capture antibody: 66444-1-PBS. Detection antibody: 60203-2-PBS. Cell lysate: Non-treated HEK293 and Calyculin A treated HEK293(30μg/well). Non-related target OAT Monoclonal Matched Antibody Pair (MP50109-1P) was served as control.

---

Recommended dilution

Application	Dilution
It is recommended that this reagent should be titrated in each testing system to obtain optimal results.

Product Information

66444-1-PBS targets Phospho-AKT (Ser473) as part of a matched antibody pair:

MP50177-1: 66444-1-PBS capture and 60203-2-PBS detection (validated in Cytometric bead array)

Unconjugated mouse monoclonal antibody pair in PBS only (BSA and azide free) storage buffer at a concentration of 1 mg/mL, ready for conjugation.

This conjugation ready format makes antibodies ideal for use in many applications including: ELISAs, multiplex assays requiring matched pairs, mass cytometry, and multiplex imaging applications.Antibody use should be optimized by the end user for each application and assay.

• Tested Reactivity: human, mouse, rat
• Host / Isotype: Mouse / IgG1
• Class: Monoclonal
• Type: Antibody
• Immunogen: Peptide
• Full Name: v-akt murine thymoma viral oncogene homolog 1
• Observed Molecular Weight: 60-62 kDa
• GenBank Accession Number: NM_005163
• Gene Symbol: AKT1
• Gene ID (NCBI): 207
• Conjugate: Unconjugated
• Form: Liquid
• Purification Method: Protein A purification
• Storage Buffer: PBS Only
• Storage Conditions: Store at -80°C.

Background Information

The serine-threonine protein kinase AKT1 is catalytically inactive in serum-starved primary and immortalized fibroblasts. Survival factors can suppress apoptosis in a transcription-independent manner by activating the serine/threonine kinase AKT1, which then phosphorylates and inactivates components of the apoptotic machinery. This antibody detects all the members of AKT with phospho-modification at Ser473.