Size-based protein separations by microchip electrophoresis using an acid-labile surfactant as a replacement for SDS.
We demonstrate the use of an acid-labile surfactant (ALS) as a replacement for SDS for size-based protein separations in a microfluidic device. ALS is of interest to the proteomic field as it degrades at low pH and hence can be removed to reduce surfactant interference with down-stream MS. A range of SDS and ALS concentrations were tested as denaturants for microchip electrophoresis to investigate their effects on the separation of proteins from 18 to 116 kDa and to provide a suitable comparison between the two surfactants. The electrophoretic mobilities of the proteins were not significantly affected by the use of ALS instead of SDS. Protein separations with ALS are performed in less than 3 min, which is a significant decrease in the time compared with the previous ALS separations on a slab gel format. We also demonstrate the use of poly-N-hydroxyethylacrylamide as a dynamic, hydrophilic chip channel coating that can be applied with a rapid and simple protocol for size-based protein separation. The results reported here could significantly decrease the time and increase the attainable level of automation and integration of the front-end protein fractionation required for "top-down" proteomics.
Root
Brian E
BE
Department of Materials Science and Engineering, Northwestern University, Evanston, IL, USA.
Zhang
Bin
B
Barron
Annelise E
AE
D046209:Electrophoresis, Microchip / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D011506:Proteins / Q000302:isolation & purification*; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D013501:Surface-Active Agents / Q000737:chemistry*
D011506:Proteins; D013501:Surface-Active Agents; D012967:Sodium Dodecyl Sulfate