Structure-performance analysis of oligo(dT) resin affinity chromatography for RNA separations

Area of Science:

• Biotechnology
• Chemical Engineering
• Materials Science

Background:

• Messenger RNA (mRNA) is a rapidly developing therapeutic and prophylactic drug modality.
• Oligo deoxythymidine (oligo(dT)) ligand chemistries are vital for mRNA chromatographic purification.
• Beaded oligo(dT) resins exhibit limitations in dynamic binding capacity (DBC).

Purpose of the Study:

• To establish a structure-performance relationship for beaded oligo(dT) resins.
• To understand the impact of resin structure on messenger RNA (mRNA) binding.
• To inform the design of next-generation resins for mRNA purification.

Main Methods:

• Surface area analysis using Brunauer-Emmett-Teller (BET) method.
• Pore size distribution and morphology characterization via mercury intrusion porosimetry and scanning electron microscopy (SEM).
• Poly-adenine RNA binding and breakthrough experiments to assess dynamic binding capacity (DBC).

Main Results:

• Resin surface areas ranged from 13-18 m²g⁻¹, with modal pore diameters between 298-526 nm.
• RNA binding occurred primarily on the bead exterior, with gradual penetration into the pore network.
• A 257% increase in DBC₁₀ was observed with increasing residence time, emphasizing diffusive transport.

Conclusions:

• Resin structural properties significantly influence messenger RNA (mRNA) binding and purification performance.
• Diffusive transport limitations impact the dynamic binding capacity (DBC) of beaded oligo(dT) resins.
• Understanding structure-performance relationships is key to optimizing resin design for efficient mRNA purification.