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Determination of Protein Phosphorylation by Polyacrylamide Gel Electrophoresis (PAGE) (CAT#: STEM-ET-0331-ZJF)

Introduction

Phosphorylation is the most important modification for protein regulation; it controls many signal transduction pathways in all organisms.<br />This service provides a polyacrylamide gel electrophoresis (PAGE) method to measure the ratio of protein phosphorylation in cells.




Principle

Polyacrylamide gel electrophoresis (PAGE) is an electrophoresis which based on polyacrylamide gel. Polyacrylamide gel is tougher than agarose gel and is relatively inert, thermostable, strong, and transparent, with a uniform pore size. PAGE is a widely used technique for separating and analyzing proteins based on their size and charge. It involves the migration of proteins through a porous gel matrix made of cross-linked polyacrylamide under the influence of an electrical field. Known as one of the most versatile and widely used techniques, PAGE has the ability to separate proteins at high resolution, allowing the detection of minor differences in protein composition.

Applications

Microbiology, molecular biology, phosphorylation, protein

Procedure

1. Preparation: Prepare electrophoresis buffer with a certain pH. Prepare a gel solution of appropriate concentration, taking care not to produce bubbles, and use a cast to solidify the solution into a gel.
2. Sample Application: Put the prepared gel with a cast into the electrophoresis tank, and add an appropriate amount of buffer. Pipette the sample into the sample wells.
3. Electrophoresis: Adjust the appropriate distance between the electrodes. Switch on the electrophoresis apparatus and set the voltage and current. Perform electrophoresis for a period of time.
4. Determination: After electrophoresis, stain the gel for visible results. Observe, record and analyze the position of separated bands. Or, utilize an imaging system for analysis.
5. Downstream processing: After separation, an additional method is often applied to the separated bands for further processing. For example, cut the band out of the gel as a slice to dissolve and purify it.

Materials

• Gel electrophoresis apparatus
• Sample solution
• Buffer solution and polyacrylamide gel