Screening Binding Conditions on CIM Oligo dT 96-Well Plate to Optimize Dynamic Binding Capacity

Buffer conditions (salt, additives) influence mRNA binding on Oligo dT. Three contributing factors were identified and tested: NaCl, MgCl2 and Gu-HCl, the latter leading to a capacity of >6 mg/mL.

Affinity-based chromatographic isolation of mRNA is robust and simple, lending itself as a useful industrial platform. mRNA constructs contain a 3’ polyA tail to increase stability in vivo, thereby affording the possibility of affinity purification using oligo-deoxythymidinic acid (Oligo dT) probes covalently coupled to a solid support. Poly-adenylated mRNA forms a stable hybrid with Oligo dT under high-salt conditions which is destabilized when the salt is removed, allowing mRNA to be released.

Due to an increasing productivity of IVT reaction, finding conditions that increase binding capacity of Oligo dT has been an intense focus of development. Multi-parallel approaches, such as screening in multi-well plate format, can significantly cut the development time by screening multiple conditions at once. 96-well plates can then be scaled-up to preparative scale, such as CIMmultus product line operated by chromatographic skids.

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