Application of Sodium Dodecyl Sulfate-Capillary Gel Electrophoresis to the Characterization of Ricin A-Chain Immunotoxins

摘要

This study was undertaken to use sodium dodecyl sulfate-capillary gel electrophoresis (SDS-CGE) for the characterization of immunotoxins constructed by cross-linking ricin A-chain (RTA) to monoclonal antibody (MAb). SDS-CGE could separate mono- and di-RTA-conjugated MAbs from unreacted MAb, as identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Resolution of RTA-MAb conjugates was improved by decreasing the running voltage from 15 to 5 kV. SDS-CGE was also useful for characterizing aggregates of RTA or deglycosylated RTA (dgRTA) that was formed during immunotoxin preparation. Of the RTA isomers with different carbohydrate contents, RTA₁, which has low carbohydrate content, was found to be more susceptible to aggregation than RTA₂. In the SDS-CGE of dgRTA, both covalently and non-covalently linked aggregates were observed under reducing and non-reducing conditions. The findings of this study suggest that SDS-CGE can be used to optimize methods of preparing effective immunotoxins and for checking degradants and aggregates.