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A Taylor Dispersion Analysis Method for the Sizing of Therapeutic Proteins and their Aggregates Using Nanolitre Sample Quantities
Published: 25 February 2011 by MDPI in The 1st Electronic Conference on Pharmaceutical Sciences session Beyond Colour - Insight into Pharmaceutical Dosage Forms by New Imaging Techniques
Abstract: Introduction: Protein aggregation and its reversibility over time has been measured for a number of protein systems by Taylor dispersion analysis (TDA). This technique has many advantages over the conventional techniques currently employed; including its fast analysis times, large concentration range, and nanolitre sample sizes. Here the results obtained by TDA are compared to those obtained from dynamic light scattering (DLS). Materials and Methods: The ActiPix D100 employs UV area imaging and Taylor dispersion analysis (TDA) for determining diffusion coefficients and hydrodynamic radii of proteins in solution. The detector monitors broadening of a band of a therapeutic protein injected into a stream of buffer solution being driven through a fused-silica capillary. The band is imaged at two windows, the first on entry to and the second on exit from a loop in the capillary. The hydrodynamic radius follows from the measured differences between peak times and variances at the two windows. This technique was used to analyse various unstressed and stressed BSA samples over periods of time. The results were compared to those obtained from DLS and thermal methods. Results and Discussion: Hydrodynamic radius results obtained using TDA were in good correlation to those obtained using DLS. TDA and DLS both indicated that over time some aggregates were resolubilised and that the aggregate level had decreased, however DLS was less sensitive than TDA to these subtle changes. TDA was also shown to be more advantageous for measuring aggregation given it is not as susceptible to large particulates affecting the results as was the case with the DLS. Conclusions: Results for these samples indicate good correlation between the techniques for investigating protein aggregation behaviour. The ability to quantify accurately aggregate levels is being explored. TDA showed notable advantages over DLS when looking at solutions containing large particulates.
Keywords: Keywords: Taylor dispersion analysis, Dynamic light scattering, Protein aggregation, nanolitre