Shreffler: SDS-PAGE

Overview

From wikipedia: "SDS-PAGE, sodium dodecyl sulfate polyacrylamide gel electrophoresis, describes a technique widely used in biochemistry, forensics, genetics and molecular biology to separate proteins according to their electrophoretic mobility (a function of the length of a polypeptide chain and its charge). In most proteins, the binding of SDS to the polypeptide chain imparts an even distribution of charge per unit mass, thereby resulting in a fractionation by approximate size during electrophoresis."

We use the Invitrogen NuPAGE system.

Materials

• Appropriate gelbox/voltage source
• NuPAGE 4-12% BIS-TRIS Gel (Invitrogen NP0322BOX)
• NuPAGE MES SDS Running Buffer (20x) (Invitrogen NP0002)
• NuPAGE SimplyBlue SafeStain (Invitrogen LC6060)
• NuPAGE LDS Sample Buffer (4X) (Invitrogen NP0007)
• NuPAGE Antioxidant (Invitrogen NP0005)
• NuPAGE Reducing Agent (Invitrogen NP0004)

Procedure

Preparing Samples

1. We want to Add 5 μL 4X Reducing Buffer (RB) to 15μL protein solution. Vortex sample to mix. Heat samples at 70 °C for 10 mins. Spin samples.

Discussion

discuss this protocol

Contact

• Who has experience with this protocol?
• Email Alex Ma through OpenWetWare