Noncooled capillary inlet: a source of systematic errors in capillary-electrophoresis-based affinity analyses

Area of Science:

• Analytical Chemistry
• Separation Science
• Biophysical Chemistry

Background:

• Capillary electrophoresis (CE) is a versatile technique for affinity-based analyses, often sensitive to temperature variations.
• Active cooling systems manage electrolyte temperature during electrophoresis, but short, uncooled sections, like the capillary inlet, exist.
• These uncooled regions have historically been overlooked as sources of experimental artifacts.

Purpose of the Study:

• To investigate the impact of uncooled capillary inlet regions on quantitative accuracy in CE-based affinity methods.
• To identify and characterize systematic errors arising from elevated temperatures in the non-cooled inlet.
• To propose a practical solution to mitigate temperature-induced artifacts in CE affinity analyses.

Main Methods:

• Experimental analysis of sample migration through the uncooled capillary inlet during CE.
• Quantitative assessment of affinity binding under varying temperature conditions at the inlet.
• Development and testing of a method to move the sample rapidly through the uncooled zone.

Main Results:

• Electrophoresis through the short, uncooled capillary inlet introduces significant systematic errors in quantitative affinity measurements.
• Elevated electrolyte temperatures in the inlet region are identified as the primary cause of these analytical artifacts.
• The magnitude of errors is substantial, impacting the reliability of CE-based affinity analyses.

Conclusions:

• The uncooled capillary inlet, despite its short length, is a critical source of artifacts in quantitative CE affinity assays.
• Developers and users of CE affinity methods must account for potential temperature-related errors originating from the inlet.
• A simple method involving pressure or low-field electrophoresis to quickly move samples into the cooled zone effectively prevents these artifacts.