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联系电话
4000-520-616 / 18915418616
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0512-67156496
电子邮箱
info@ebiomall.com
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https://www.ebiomall.com
商品描述
Antigen Information
Accession Number:
- P31749
Gene ID:
- 207
Gene Symbols:
- AKT1
- PKB
- RAC
Protein Name & Synonyms:
RAC-alpha serine/threonine-protein kinase (EC 2.7.11.1) (Protein kinase B) (PKB) (Protein kinase B alpha) (PKB alpha) (Proto-oncogene c-Akt) (RAC-PK-alpha)
Assay Format
Pathway:
- Insulin Signaling
- mTOR Signaling
- PI3K-AKT Signaling
Specificity:
The antibodies provided in this kit recognizes human/mouse/rat AKT phosphorylated at Serine-473 and total AKT for comparison.
Number of Targets Detected:
1
Species Detected:
- Human
- Mouse
- Rat
Compatible Sample Types:
- Adherent Cell Culture
Design Principle:
- Cell-based
Method of Detection:
Colorimetric
Quantitative/Semi-Quantitative:
- Semi-Quantitative
Solid Support:
96-well Microplate
Product Specifications
Size:
1, 2, or 5 x 96-Well Microplate Kit
Product Features
- Site and signal pathway-specific
- In vitro detection of adherent cell culture
- No sample lysis is needed
- Compatible with a standard ELISA plate reader
- Faster results than with ELISA
- Adaptable for high-throughput screening and drug discovery
Application Notes
Kit 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
Other Materials Required
- 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
Protocol Outline
- Seed 10,000-30,000 cells into each well and incubate overnight.
- Apply various treatment, inhibitors or activators according to manufacture"s instructions.
- Add 100 µl of Fixing Solution into each well and incubate for 20 min at RT with shaking.
- Add 200 µl of prepared 1X Quenching Buffer and incubate 20 min at RT.
- Add 200 µl of Blocking Solution and incubate for 1 h at 37 °C.
- 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.
- Add 50 µl of prepared 1X HRP-Anti-Rabbit or Mouse IgG and incubate for 1 h at RT.
- Add 100 µl of TMB One-Step Substrate Reagent to each well.
- Incubate 30 min at RT.
- Add 50 µl of Stop Solution to each well.
- Read at 450 nm immediately.
Storage/Stability
The entire kit may be stored at -20°C for up to 6 months from the date of shipment. Avoid repeated freeze-thaw cycles.
SUNY Stony Brook University06/12/2018, 11:23 AM
Manual
SDS
