Taylor Dispersion Analysis (TDA) is performed using capillaries with a small inner diameter. However, the strong tendency of proteins to adhere to the capillary walls can distort peaks, reduce reproducibility, and compromise accuracy. In this study, we propose a simple yet effective approach to minimize interactions between proteins and the capillary wall. When TDA is carried out rapidly, the leading portion of the parabolic flow profile avoids contact with the wall, allowing a significant fraction of the sample to remain in the center of the capillary. As a result, large molecules at the front of the injected sample plug experience minimal or no adsorption. We examined both the theoretical basis and practical implementation of this fast TDA approach, where the residence time is shorter than 1 min. Our findings indicate that diffusion coefficients of proteins should be determined using different strategies depending on the residence time: (i) measuring twice the front half-width for standard TDA, (ii) fitting a Gaussian curve for fast TDA with a 0.7 min residence time, or (iii) employing a calibration curve (w25% vs known diffusion constants) for ultrafast TDA with a 0.07 min residence time. That is, in some cases of rapid analysis, the standard equation used in TDA for absolute diffusion coefficient determination may carry significant error; thus, the use of a calibration-based method is suggested in such instances. In general, diffusion coefficients obtained via fast TDA tend to be slightly larger than those obtained from conventional TDA. This can be explained by the reduction of peak broadening effects caused by protein adsorption achieved by the fast TDA approach.
Fast Taylor Dispersion Analysis for Minimizing Protein Adsorption Effects
Subjects: Proteins