Successful commercialization of RNA therapeutics will require the development of appropriate manufacturing and purification processes suitable for the production of large quantities of high quality and high purity RNA with well-defined tertiary structures. Ultrafiltration has the potential to play a major role in RNA processing, both for removal of large molecular weight species (with the RNA collected in the permeate) and for RNA concentration or buffer exchange (with the RNA collected in the retentate). Previous studies have demonstrated the potential of using tangential flow filtration (TFF) for RNA separations, but most of these data were obtained with crude RNA mixtures containing RNA with a range of size and structure. The objective of this study was to directly evaluate the effects of RNA size and tertiary structure on the ultrafiltration behavior over a range of solution conditions (e.g., added salt and urea). Transmission of a hairpin RNA with 70 nucleotides was much greater than that of a linear RNA having the same number of nucleotides, consistent with the smaller size as determined by dynamic light scattering. RNA transmission increased at high NaCl concentrations and in the presence of urea, particularly for the linear RNA. These results provide new insights into RNA ultrafiltration as well as a framework for the application of ultrafiltration for the purification of RNA therapeutics.
All Science Journal Classification (ASJC) codes
- Analytical Chemistry
- Filtration and Separation