Tuning Apparent Peak Efficiency in Capillary Electrophoresis Using Backscatter Interferometry Detection

Backscatter interferometry (BSI) is a refractive index detection method for capillary electrophoresis that is inexpensive, flexible, and easily miniaturized. Interestingly, unlike most detectors that respond exclusively to analyte concentration, the BSI signal is sensitive to both refractive index (analyte concentration) and the separation voltage. The latter is linked to zone conductivity and leads to improved BSI signals and lower detection limits with increasing field strengths. Enhanced BSI signals can also be generated using a photothermal mechanism, where resonantly excited analytes release heat into their surroundings to increase the BSI signal amplitude. Both voltage-based and photothermal signal enhancement mechanisms can lead to a change in the polarity of the BSI signal, which can be either positive or negative depending on the specific analyte, its concentration, and the separation conditions. Here, we show that this leads to a significant increase in apparent peak efficiency. At the transition in peak polarity, both mechanisms result in over a 10-fold increase in apparent peak efficiency, improving from approximately 10⁵ plates/m to over a million plates/m. Simultaneously measured BSI and fluorescence electropherograms confirm that the efficiency increase is unique to the BSI signal and not due to changes in zone dispersion, and can be tuned to optimize separation resolution. The origin of the efficiency increase is discussed in terms of the refractive index and zone conductivity contributions to the BSI signal.