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Hyaluronan Molecular Mass Determination by Agarose Gel Electrophoresis (CAT#: STEM-ET-0324-ZJF)

Introduction

The average molecular mass of hyaluronan (HA) in most healthy biological fluids and tissues is usually about 6000–8000 kDa, but the biosynthetic mechanism results in a polydisperse mixture of sizes. Subsequent enzymatic degradation, or the action of reactive oxygen and nitrogen species, can further increase polydispersity and decrease the average size. Fragmented HA can be a biomarker of inflammation. In addition, reductions in HA size are associated with tissue remodeling and repair processes. This service provides an agarose gel electrophoresis method for the determination of the molecular mass distribution of HA isolated from tissues and fluids.




Principle

Gel Electrophoresis is one of the zone electrophoresis, and the supporting medium of it is gel. The gel is a cross-linked polymer, acting as a molecular sieve. Its composition and porosity are chosen based on specific requirements. The types of gel typically used are starch, agarose, and polyacrylamide gels.
Agarose is a purified uncharged polysaccharide extracted from seaweeds. By manipulating the concentration of agarose in the gel, the pore size can be adjusted. Agarose gel electrophoresis is suitable for the separation of proteins larger than 200 kDa, and DNA fragments of 0.2-20kb.

Applications

Molecular biology, hyaluronan

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 agarose gel
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