Mechanism of Protein Isoform Analysis by CE-SDS

Proteins are critical molecules in living organisms, performing various cellular functions. Protein isoform analysis is an essential method for studying protein heterogeneity. Capillary Electrophoresis-Sodium Dodecyl Sulfate (CE-SDS) is a widely used analytical technique for protein isoform analysis.

 

Basic Principles of CE-SDS

The working principle of CE-SDS is based on the binding of SDS to proteins, forming negatively charged protein-SDS complexes. Under the influence of an electric field, these complexes migrate within the capillary and are separated based on their molecular weight. This process includes the following steps:

 

1. Preparation of Protein Samples

Samples must first be denatured with SDS and a reducing agent (such as dithiothreitol) to break disulfide bonds and secondary structures, allowing the proteins to fully unfold and bind with SDS.

 

2. Capillary Preparation

The capillaries are typically coated with a non-polar substance like polyacrylamide to prevent protein adsorption onto the inner walls of the capillary, ensuring smooth migration of protein-SDS complexes.

 

3. Electrophoretic Separation

Under the electric field, protein-SDS complexes are separated based on their molecular weights. Smaller molecules migrate faster, while larger molecules migrate slower, achieving effective separation within the capillary.

 

Data Analysis and Interpretation

CE-SDS results are usually presented as electropherograms, where each peak represents a protein isoform. By comparing with standard proteins, the molecular weights and relative quantities of different protein isoforms in the sample can be determined.

 

Applications

CE-SDS is widely applied in biopharmaceuticals. For example, during monoclonal antibody production, CE-SDS is used to detect antibody purity and heterogeneity. Additionally, CE-SDS is employed in vaccine development and quality control to ensure vaccine safety and efficacy.

 

MtoZ Biolabs provides integrate CE-SDS protein isoform analysis service.