Capillary Gradient Gel Electrophoresis

In the last half-century, capillary gel electrophoresis (CGE) became a versatile and high-performance analytical platform for the separation of complex biomolecular mixtures featuring rapid separations, high efficiency, and small sample consumption. Integrating a pore-size gradient mechanism in CGE makes it possible to achieve enhanced selectivity of polyionic macromolecules such as SDS-proteins and nucleic acids. This review provides a comprehensive overview of the theoretical foundations and operational principles of capillary pore-size gradient gel electrophoresis (CGGE), including the physicochemical basis of gradient formation, the influence of pore-size distributions on analyte mobility, and the challenges of generating stable, reproducible gradients in narrow-bore capillaries. Instrumental considerations such as capillary surface treatment, gradient filling and polymerization strategies, temperature and voltage control, detection modalities, and method-development frameworks are discussed in detail, emphasizing their critical impact on analytical performance and reproducibility. Key application areas in bioanalytical chemistry are highlighted, covering nucleic acid analysis and peptide/protein characterization. CGGE offers unique analytical advantages where fine molecular discrimination, tunable selectivity, and high resolution in a broad molecular weight range are required.