SuperSepTM Phos-tagTM (50μmol/L), 15%, 17well
- for Electrophoresis
- Manufacturer :
- FUJIFILM Wako Pure Chemical Corporation
- Storage Condition :
- Keep at 2-10 degrees C.
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- SDS
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Document
Applications
Albumin Dephosphorylation
[Electrophoresis Buffer] Tris-Glycine SDS Buffer
[Electrophoresis Samples]
Lane1: Untreated Albumin
Lane2: Dephosphorylated Albumin
[Electrophoresis cConditions] 20 mA, 70 mins
[Staining] Quick CBB staining
[Destaining] Deionized wWater
Albumin (Cat. No. 010-17071) was dephosphorylated using alkaline phosphatase (NIPPON GENE CO., LTD., Cat. No. 319-02661). Band shift confirmed dephosphorylation of albumin.
Time-dependent Dephosphorylation of β-casein
[Electrophoresis Buffer] Tris-Glycine SDS buffer
[Electrophoresis Samples]
Lane1: β-casein (AP-treatment: 0 min)
Lane2: β-casein (AP-treatment: 15 min)
Lane3: β-casein (AP-treatment: 30 min)
Lane4: β-casein (AP-treatment: 45 min)
Lane5: β-casein (AP-treatment: 60 min)
[Electrophoresis Conditions] 35mA, 60min
[Staining] Quick CBB staining
[Destaining] Deionized Water
Dephosphorylation of β-casein was performed over different time periods. The results confirmed separation of phosphorylated and dephosphorylated β-casein.
In addition, the results demonstrated the level of dephosphorylation across different time points.
Time-dependent Change in the level of phosphorylation
Procedure
X-ray irradiation (5 Gy) was performed for human lung cancer cell line Lu99, and cells were collected across different time points. Cell extracts were prepared, and SDS-PAGE was performed in 13 wells using 50 μmol/L of 10% SuperSep™ Phos-tag™
Gentle agitation of the gel was performed in a transfer buffer containing 10 mmol/L EDTA, and proteins were transferred to PVDF membrane. The membrane was blocked with 2% Milk/TBS-T, and was reacted with primary antibodies (upper image: p53, lower image: cell-cycle proteins). Detection was performed by chemoluminecence.
Results
Protein accumulation for p53 was at the highest level 4 hours after X-ray irradiation. The level of protein X phosphorylation changed over time after X-ray irradiation.
Electrophoresis of Control Protein
P10 %(left): SuperSep™Phos-tag™ (50 μmol/L), 10 %, 13Wells
P15 %(middle): SuperSep™Phos-tag™ (50 μmol/L), 15 %, 13Wells
A15 %(right): SuperSep™Ace, 15 %, 13Wells
Electrophoresis Conditions
30 mA/gel (constant), 60 mins
Sample
5 μg/lane of α-casein from bovine milk, dephosphorylated (Cat. No. 038-23221)
- The product contains α-casein that hasve not been dephosphorylated. While conventional SDS-PAGE will only show a single main band, Phos-tag™ SDS-PAGE will show two main bands for α-casein and dephosphorylated α-casein.
Running Buffer
Staining
Quick CBB Plus (Cat. No. 174-00553)
Overview / Applications
| Outline | This product is for research use only. Do not administer it to human.
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Alias
For research use or further manufacturing use only. Not for use in diagnostic procedures.
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