ELECTROPHORESIS: THEORY AND APPLICATIONS

Electrophoresis serves as a foundational analytical technique supporting modern molecular biology, biochemistry, clinical diagnostics, forensic science, and biotechnology. Originating in the early nineteenth century with the discovery of electrokinetic particle migration in electric fields, electrophoresis has experienced ongoing theoretical and technological advancements. This chapter provides a comprehensive and integrative overview of electrophoresis, including its historical development, fundamental physicochemical principles, supporting media, buffer systems, and principal methodological variants that characterize current practice. The chapter initially outlines key milestones in the development of electrophoresis, ranging from early electrochemical observations and moving-boundary electrophoresis to the introduction of zone electrophoresis, gel-based separations, and high-resolution capillary electrophoresis. It examines the fundamental principles governing electrophoretic migration, including the influence of charge, electric field strength, electrophoretic mobility, viscous drag, steric hindrance, and properties of the separation medium. Supporting matrices and buffer systems are evaluated as primary determinants of separation efficiency. The chapter addresses free-solution electrophoresis, paper and cellulose acetate systems, agarose gels, and polyacrylamide gels, with attention to their structural features, sieving properties, and analytical applications. Major electrophoretic techniques are systematically reviewed, including slab (zone) electrophoresis, nucleic acid and protein gel electrophoresis, capillary electrophoresis and its variants, high-voltage systems, pulsed-field gel electrophoresis, and specialized protein methods such as SDS–PAGE, native and blue-native PAGE, isoelectric focusing, and two-dimensional electrophoresis. Recent developments in microfluidic and lab-on-a-chip electrophoresis are highlighted as transformative innovations that facilitate rapid, automated, and low-volume analyses with potential for point-of-care applications. The chapter concludes by addressing current limitations and future directions, with particular emphasis on integration with mass spectrometry, sequencing technologies, automation, and data-driven analytical frameworks. Overall, electrophoresis is presented as a dynamic and evolving methodology that remains essential for high-resolution molecular characterization in both research and applied biomedical sciences.