Principle
In capillary electrophoresis (CE), an electric field acts on dissolved charged species inside a narrow capillary containing a background electrolyte. Species separate when their electrophoretic mobilities differ. Mobility depends on charge, effective size and shape, solvation, temperature, and the properties of the surrounding liquid.
Charged capillary walls can also generate electroosmotic flow (EOF), which moves the liquid as a whole. An analyte’s observed velocity is the combination of its electrophoretic motion and EOF. Those contributions may reinforce or oppose one another, so EOF should not be assumed to point toward the detector under every surface treatment, electrolyte, or polarity.
A terminology boundary worth knowing
“CE” is often used informally for several techniques run in capillaries or on related instruments. In strict IUPAC usage, CE denotes separation based on electrophoretic-mobility differences. Micellar electrokinetic chromatography (MEKC) adds retention in a moving pseudostationary phase, while capillary electrochromatography (CEC) is a liquid-chromatographic technique whose mobile phase is commonly driven by EOF. The broader phrase capillary electromigration techniques avoids hiding these different mechanisms.
Instrumentation and workflow
A basic CE system includes two electrolyte reservoirs, a capillary, a high-voltage supply, a way to introduce a narrow sample zone, and a detector. Commercial systems commonly add pressure or vacuum control, temperature regulation, automated rinsing, vial handling, and data processing. An electropherogram plots detector response against migration time.
Capillary surface chemistry, electrolyte composition and pH, ionic strength, injection mode, field strength, temperature, and detector geometry all affect the result. Hydrodynamic and electrokinetic injections can introduce different biases, especially when sample conductivity varies.
CE and related capillary techniques
- Capillary zone electrophoresis (CZE): free-solution separation governed mainly by differences in electrophoretic mobility.
- Capillary gel electrophoresis (CGE): a polymer network or gel adds molecular sieving, widely used for nucleic acids and proteins.
- Capillary isoelectric focusing (CIEF): amphoteric species focus where the local pH matches their isoelectric point; the result is a focused spatial pattern rather than ordinary zone migration.
- Micellar electrokinetic chromatography (MEKC): analytes distribute differently between the surrounding solution and micelles that act as a moving pseudostationary phase, permitting separation of neutral as well as charged compounds.
- Capillary electrochromatography (CEC): chromatographic retention occurs in a capillary containing a stationary phase, while EOF commonly drives the liquid mobile phase.
Strengths
- very small sample consumption and low reagent use;
- high separation efficiency in appropriate systems;
- electronic control with few moving fluidic parts;
- several complementary mechanisms in related capillary instrument architectures;
- natural connection to microchip formats and miniaturization.
Limitations and interpretation traps
- migration time is conditional evidence, not a universal molecular identity;
- adsorption to capillary walls can distort peaks and recovery;
- small optical path lengths can limit concentration sensitivity in absorbance detection;
- low sample consumption does not by itself guarantee a low concentration detection limit;
- Joule heating, electrolysis-driven changes in reservoir composition or pH, and injection bias can undermine performance;
- neutral analytes have no electrophoretic mobility and require another basis for separation, such as MEKC or CEC;
- coupling to mass spectrometry introduces interface and electrolyte constraints.
Detection and coupling
UV/visible absorbance, laser-induced fluorescence, conductivity, electrochemical detection, and mass spectrometry are common detector families. The detector changes which analytes are observable, how much sample preparation or labeling is needed, and whether a compact format remains practical.
Where CE is especially useful
CE and neighboring capillary electromigration techniques are used for species ranging from small inorganic ions to proteins, therapeutic biomolecules, nucleic acids, particles, and cells. They are especially attractive when sample is scarce, charge-related selectivity matters, or low-volume electrical control is valuable.