Collection: Capillary Electrophoresis (CE) Chemicals and Accessories

Capillary Electrophoresis

In the realm of Capillary Electrophoresis (CE) and related techniques, capillaries are almost exclusively crafted from drawn fused silica, bolstered with an external polyimide coating to fortify their strength and mechanical stability. Typically, the inner walls of these capillaries are left uncoated, but specific substances can be bonded to the inner surface for two primary purposes: to eliminate undesirable interactions between the sample and the wall or to modify the extent and direction of electro-osmotic flow (EOF).

Bare Silica Capillaries

Inside bare silica capillaries, the inner wall boasts numerous surface silanol groups. When these groups come into contact with an electrolyte solution, they dissociate, resulting in a negative charge on the capillary wall. To maintain overall electrical neutrality, there must be an excess of cations in the electrolyte immediately adjacent to the inner surface. When a voltage is applied, these cations migrate towards the cathode, inducing a flow in the same direction known as electro-osmotic flow (EOF). In bare silica capillaries (uncoated silica), the EOF is particularly pronounced when the pH of the electrolyte exceeds 6, and it effectively diminishes to zero for pH values of 3 or less.

For consistent and reproducible analyses, it is crucial that the surface density of free silanol groups remains constant from one analysis to the next. To achieve this, capillaries are often rinsed with sodium hydroxide solution (or a strong acid) followed by a further rinse with the running buffer. In the case of coated capillaries, the rinsing procedures are typically less harsh to avoid stripping off the inner coating.

To ensure a uniform EOF, new uncoated capillaries are initially flushed with a 0.1M NaOH solution for 20 minutes. This process reactivates the surface silanols. It's important to note that this procedure should not be performed with wall-coated capillaries.

Electrical Double Layer - Origin of Electro-Osmotic Flow (EOF)

Electrical Double Layer - Origin of Electro-Osmotic Flow (EOF)

Capillary Regeneration

Capillary Regeneration

Coated Capillaries

The composition of the capillary's inner wall can be altered through covalent bonding with suitable substances. This modification may result in a neutral or charged surface layer.

Neutral Coatings

Neutral capillaries serve to reduce analyte adsorption or eliminate EOF. Typically, these capillaries are covalently coated with polymers like celluloses or polyacrylamide, effectively eradicating EOF. Before the bonding process, the silica surface is typically derivatized with an alkyl silane.

Charged Coatings

Capillaries can also be coated with positively or negatively charged substances to manipulate the direction and magnitude of EOF and/or reduce analyte adsorption. For instance, polyamine-coated capillaries create a highly positively charged surface, leading to a substantial EOF towards the anode—a departure from the usual direction. This positive charge repels positively charged basic compounds, reducing their tendency to adsorb on the surface.

Conversely, capillaries can be treated to enhance the negative charge on the surface compared to an uncoated capillary. This increased EOF can be advantageous in significantly reducing analysis times.

Unlike uncoated capillaries, where EOF strongly depends on the buffer pH and is absent at low pH levels, a fully ionized capillary coating over a broad pH range results in an EOF rate largely independent of pH (pH-independent capillary). This feature allows for the utilization of surfactant-based micellar electrokinetic chromatography (MECC) at lower pH values than typically employed. Low pH MECC is particularly valuable for analyzing acidic solutes that are negatively charged at high pH levels and therefore repelled by negatively charged micelles.

Precise control of surface charge density on pH-independent coatings enables fine-tuning of EOF. A medium charge density produces a moderate EOF rate (medium flow capillary), while a low level of coating results in a low EOF (low flow capillary).

Additional Information on Capillaries

Uncoated CE capillaries are constructed from high-quality drawn fused silica, ensuring excellent UV transparency. All Capital Analytical capillaries possess an outer diameter of 375μm and are available with internal diameters of 50μm and 75μm.

For ease of handling and installation, most CE instruments utilize a cartridge that houses the coiled capillary. Standard capillary lengths are typically 25-75cm, with longer lengths, such as 100cm or more, occasionally employed for specialized applications, like interfacing commercial CE instruments with mass spectrometers.

Replacing capillaries can be tricky since the removal of the polyimide coating to create the detection window leaves the exposed fused silica quite fragile. To mitigate the risk of damage, Capital Analytical offers prepared capillaries that are pre-fitted in cartridges, precisely aligned with the detection aperture to ensure maximum UV sensitivity. Proper cutting of the injection end of the capillary is crucial for maintaining peak precision and preventing issues like sample carry-over. Capital Analytical's prepared capillaries are expertly cut with a precisely positioned UV detection window and a portion of the polyimide coating removed from the injection end to enhance sample introduction accuracy.

Additionally, Capital Analytical can provide capillaries in non-standard lengths or offer precision preparation services for customers who prefer to supply their own capillaries. The company also offers capillary replacement services for various cartridge formats.

For inquiries about capillary replacement for specific instrument formats, please contact our sales office.