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Isoelectric Focusing Array with Immobilized pH Gradient and Dynamic Scanning Imaging for Diabetes Diagnosis (CAT#: STEM-ET-0358-ZJF)

Introduction

As one of the most efficient separation techniques, isoelectric focusing (IEF) has been widely used in 2D separation for the proteomics research, capillary IEF (cIEF) for the quality control of protein drugs, free-flow electrophoresis for the fractionation of cells and organelles, and gel slab IEF for the focusing of hemoglobin A1c (HbA1c). A traditional immobilized pH gradient (IPG) has a high stability for isoelectric focusing (IEF) but suffers from time-consuming rehydration, focusing and staining-imaging as well as complex performance. This service provides an isoelectric focusing method using an array of 24 IPG columns (10 mm × 600 μm × 50 μm) and dynamic scanning imaging (DSI) for protein focusing. The IEF array is applicable to diabetes diagnosis.




Principle

Isoelectric focusing (IEF) is a method of electrophoresis that separates proteins according to their isoelectric point. It performs in a particular pH gradient, in which proteins are separated by differences in their isoelectric points. The isoelectric point (pI) is the pH at which the net charge of the protein is zero. Protein is positively charged when its pI is above the solution's pH, so it migrates towards the cathode. Conversely, protein is negatively charged when its pI is below the solution's pH, so it migrates towards the anode. Protein has no net charge at its pl and stops migrating. Based on this principle, proteins can be separated into sharp bands with each protein positioned at a point in the pH gradient corresponding to its pI.

Applications

Clinical applications

Procedure

1. Preparation: Prepare the gel containing the sample. Mix the solution and add it to a glass tube to form a gel. Prepare the fixing solution, dyeing solution and decolorization solution.
2. Electrophoresis: Add the appropriate acidic solution in the positive electrophoresis tank, and the appropriate alkaline solution in the negative electrophoresis tank. Switch on the electrophoresis apparatus and set the voltage and current. Perform pre-electrophoresis for a period of time to form a pH gradient in the gel. Then set the electrophoresis apparatus to a higher voltage and perform electrophoresis for a period of time.
3. Determination: Different protein components focus on the corresponding pI. Fix, stain and decolorize the gel. Finally, dry it for storage.

Materials

• Isoelectric focusing apparatus
• Sample solution