AKT1 ELISA Kit

Catalog No. ABIN4881275

Product Details AKT1 ELISA Kit

Target: AKT1 (V-Akt Murine Thymoma Viral Oncogene Homolog 1 (AKT1))

Binding Specificity: pSer473

Reactivity: Human, Mouse, Rat

Detection Method: Colorimetric

Method Type: Cell ELISA

Application: ELISA

Purpose: Cell-Based Human/Mouse/Rat AKT (S473) Phosphorylation ELISA Kit. Suitable for adherent whole cell lines.

Sample Type: Cell Culture Cells

Analytical Method: Semi-Quantitative

Specificity: The antibodies provided in this kit recognizes human/mouse/rat AKT phosphorylated at Serine-473 and total AKT for comparison.

Characteristics:

• Site and signal pathway-specific
• In vitro detection of adherent cell culture
• No sample lysis needed
• Compatible with a standard ELISA plate reader
• Faster results than with ELISA
• Adaptable for high-throughput screening and drug discovery

Components:

• uncoated 96-well Microplate
• Wash Buffer A
• Wash Buffer B
• Fixing Solution
• Quenching Buffer
• Blocking Buffer
• Anti-phospho antibody
• Anti-pan antibody
• HRP-Conjugated Secondary Antibody
• TMB One-Step Substrate
• Stop Solution

Material not included:

• Distilled or deionized water
• 100 mL and 1 liter graduated cylinders
• Tubes to prepare sample dilutions
• Protease and Phosphatase inhibitors
• Precision pipettes to deliver 2 μL to 1 mL volumes
• Adjustable 1-25 mL pipettes for reagent preparation
• Benchtop rocker or shaker
• Microplate reader capable of measuring absorbance at 450 nm

Application Details

Sample Volume: 100 μL

Plate: Uncoated

Protocol:

1. Seed 10,000-30,000 cells into each well and incubate overnight.
2. Apply various treatment, inhibitors or activators according to manufacture's instructions.
3. Add 100 μL of Fixing Solution into each well and incubate for 20 min at RT with shaking.
4. Add 200 μL of prepared 1X Quenching Buffer and incubate 20 min at RT.
5. Add 200 μL of Blocking Solution and incubate for 1 h at 37 °C.
6. Add 50 μL of 1X anti-phospho-protein specific antibody or anti-pan-protein specific antibody to each well and incubate for 2 h at RT.
7. Add 50 μL of prepared 1X HRP-Anti-Rabbit or Mouse IgG and incubate for 1 h at RT.
8. Add 100 μL of TMB One-Step Substrate Reagent to each well.
9. Incubate 30 min at RT.
10. Add 50 μL of Stop Solution to each well.
11. Read at 450 nm immediately.

Restrictions: For Research Use only

Handling

Storage: -20 °C

Storage Comment: The entire kit may be stored at -20°C for up to 6 months from the date of shipment. Avoid repeated freeze-thaw cycles.

Expiry Date: 6 months

Target Details for AKT1

Target: AKT1 (V-Akt Murine Thymoma Viral Oncogene Homolog 1 (AKT1))

Alternative Name: Akt (AKT1 Products)

Gene ID: 207

UniProt: P31749

Pathways: PI3K-Akt Signaling, RTK Signaling, TCR Signaling, AMPK Signaling, Interferon-gamma Pathway, TLR Signaling, Fc-epsilon Receptor Signaling Pathway, EGFR Signaling Pathway, Neurotrophin Signaling Pathway, Response to Water Deprivation, Regulation of Actin Filament Polymerization, Carbohydrate Homeostasis, Glycosaminoglycan Metabolic Process, Cellular Glucan Metabolic Process, Regulation of Muscle Cell Differentiation, Cell-Cell Junction Organization, Regulation of Cell Size, Skeletal Muscle Fiber Development, Regulation of Carbohydrate Metabolic Process, Hepatitis C, Protein targeting to Nucleus, CXCR4-mediated Signaling Events, Signaling Events mediated by VEGFR1 and VEGFR2, Negative Regulation of intrinsic apoptotic Signaling, Thromboxane A2 Receptor Signaling, Signaling of Hepatocyte Growth Factor Receptor, Positive Regulation of fat Cell Differentiation, VEGFR1 Specific Signals, VEGF Signaling, Warburg Effect