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Analysis of Protein Aggregates Molecular Weight by Capillary Gel Electrophoresis (cGE) (CAT#: STEM-B-0300-CJ)

Introduction

The generic term 'aggregates' refers to species characterized by a wide size range, diverse morphologies and structures. Protein aggregates may start in the low nanometer size range but then can grow into the micrometer and even visible size range.<br /><br />Most protein therapeutics and many other biopharmaceutical compounds are inherently unstable and can undergo aggregation through various pathways. Aggregates of various kinds can be formed, such as reversible and non-reversible, soluble, and non-soluble etc. In addition,Aggregation maybe occur because of exposure to air-liquid or liquid-solid interfaces, e.g., during mixing, during filling and shipping, during reconstitution of lyophilized products, or through contact with chromatography columns, pumps, pipes, vessels, filters, etc. Aggregation can directly influence the efficacy of the therapy by reducing the number of functional molecules, but also indirectly influence efficacy as well as safety of a therapy by inducing side-effects, such as unwanted immunogenicity.<br /><br />Molecular weight, also called molecular mass, mass of a molecule of a substance, based on 12 as the atomic weight of carbon-15. It is calculated in practice by summing the atomic weights of the atoms making up the substance's molecular formula.




Principle

cGE is a high-resolution and automated variation of the SDS-PAGE technique. In general, electrophoresis is a separation technique based on the migration of charged molecules in response to an electric field, toward the electrode of opposite charge. It is performed mainly in polyacrylamide gels. In cGE, the gel is located inside a capillary through which the sample components must migrate. The larger the molecular weight, the longer the migration time. A detection system, mostly UV or fluorescence detection, detects and quantifies the migrating species. Molecular weight is determined in reference to a standard.

Applications

Biopharmaceutica

Procedure

1. The start and end vials and the connecting capillaries are filled with electrolyte solution.
2. The sample to be separated is introduced in the capillary.
3. Electric field is applied and the analytes migrate in the opposite charge direction.
4. Samples and separations are detected via various modes depending on the experimental setup.

Materials

• Sample: Proteins
• Equipment: Capillary gel electrophoresis (cGE)

Notes

cGE allows for a much higher resolution and faster sample analysis and has a lower limit of detection.