Gel electrophoresis: SDS-PAGE and native PAGE
Coriolis Pharma is a leading contract laboratory providing analytical services of protein aggregation testing, including SDS-PAGE and native PAGE analysis.
Electrophoresis is a separation method 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, which act as size-selective sieves. The most common polyacrylamide gel electrophoresis (PAGE) technique employs the detergent sodium dodecyl sulfate (SDS). SDS denatures proteins and binds those noncovalently providing an overall negative charge. Since the bound SDS/protein (w/w) ratio is practically protein independent, under a given electric field SDS-bound proteins migrate through a gel depending primarily on their size, enabling molecular weight estimation. Moreover, by comparing electrophoretic profiles of proteins analyzed under reducing and nonreducing conditions, information about the nature of covalent bonds (disulfide mediated or not) can be obtained.
SDS-PAGE allows for the qualitative and (semi-)quantitative analysis of (covalent) aggregates and fragments. Noncovalent aggregates are usually not detected, because SDS dissociates noncovalent bonds.
Native PAGE, however, can provide information about noncovalent aggregation, since electrophoresis is performed in the absence of denaturing agents. In native gels, proteins generally retain their native conformation and their mobility is governed by the ratio of electric charge to hydrodynamic size.