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SuperSepTM Phos-tagTM (50μmol/L), 12.5%, 13well

for Electrophoresis
Manufacturer :
FUJIFILM Wako Pure Chemical Corporation
Storage Condition :
Keep at 2-10 degrees C.
  • Structural Formula
  • Label
  • Packing
SDS
Comparison
Product Number
Package Size
Price
Inventory
Distributor
195-16391
Barcode No
4987481627536
5Gels
List Price
330.00 USD

In stock in Japan

Document

SDS
Product Specification Sheet
Spectral Data
Certificate of Analysis
Calibration Certificate

Features

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  • Ready-to-use, saves time
  • Able to isolate phosphorylated proteins by the level of phosphorylation
  • Good separation with sharp bands

Applications

Albumin Dephosphorylation

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[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

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[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

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Data was provided by courtesy of Dr. Atsushi Enomoto, Molecular Radiology / Sec. of Radiation Biology, Center for Disease Biology and Integrative Medicine, Faculty of Medicine, University of Tokyo
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

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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.

This is a precast gel added with Phos-tag™ Acrylamide in advance. It can be used immediately after opening the package. It contains zinc as a supplemental metal. It has excellent storage stability by its neutral gel buffer. Sharp bands can be obtained.

Features:

  1. Phosphorylated- and non-phonphorylated proteins are efficiently separable with sharp bands.

  2. Long term stability (stable for 6 months)

Physical Properties:
  • Plate size: 100 x 100 x 3 (mm)

  • Gel size: 90 x 85 x 1 (mm)

  • Well number: 13

  • Well volume: 30 μL

  • Note:
    Please note that prestain marker does not reflect the molecular weight, when a gel containing Phos-tag™ is use. It can only be used as reference of electrophoresis. Please use the result as an index of transcription efficiency at Western blotting. At Western blotting, Zn2+ must be sufficiently removed with transfer buffer containing 10 mM EDTA. Please wash out the gel for 10 minutes x 3 times. Subsequently, perform the same procedure for 10 minutes using transfer buffer without EDTA, then transfer the gel.


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