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Highly Sensitive SDS Capillary Gel Electrophoresis with Sample Stacking Requiring Only Nanograms of Adeno-Associated Virus Capsid Proteins

  • Chao-Xuan ZhangEmail author
  • Michael M. Meagher
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1972)

Abstract

Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) has been the method of choice in the past decades for size-based protein analysis. However, in general it requires the protein concentration in mg/mL level and thus is not practical for trace level protein analysis, not to mention the lengthy labor-intensive procedures. The SDS capillary gel electrophoresis (SDS CGE) method reported herein requires only nanogram-sized proteins loaded onto the autosampler. A sample stacking technique (e.g., head-column field-amplified sample stacking (HC FASS)) was employed, providing three orders of magnitude sensitivity enhancement compared to conventional SDS CGE. This method has been used routinely in purity analysis and characterization of adeno-associated virus (AAV) intermediates and finished gene therapeutics of AAV vectors. The sensitivity achieved is comparable to the currently most sensitive size-based protein assay silver-stained SDS PAGE. The highly sensitive sample stacking SDS CGE can be used for other types of proteins as well.

Key words

SDS capillary gel electrophoresis (SDS CGE) Head-column field-amplified sample stacking FASS Adeno-associated virus (AAV) protein High sensitivity 

Notes

Acknowledgements

Dr. Timothy Lockey, Dr. Bryan Piras, Dr. Jason Drury, Dr. Susan Sleep, and Dr. Clifford Froelich are greatly acknowledged for providing AAV samples and analytical support. This work was supported by American Lebanese Syrian Associated Charities (ALSAC).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Department of Therapeutics Production and QualitySt. Jude Children’s Research HospitalMemphisUSA

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