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Optimizing IVT Kinetics with Rapid HPLC Analysis using an Analytical Monolith Column

BIA-Separations (now part of Sartorius AG) has published a new poster illustrating how their analytical monolith, the CIMac™ PrimaS HPLC column, can be used for the fast analysis of an IVT (in vitro transcription) reaction.

IVT is a way to obtain highly pure and uniform mRNA oligonucleotides of lengths ranging from about 15 to several thousand nucleotides. It is transcribed from a double-stranded DNA template by RNA polymerase. The IVT reaction is one of the most expensive steps in the mRNA production process and its optimization to reach high mRNA yield is of key importance. Standard mRNA quantification techniques like absorbance and fluorescence based assays are time consuming and cannot be performed at line as the IVT reaction progresses. In addition, other reaction components like nucleotides and pDNA interfere in the analytical results and reduce the method’s accuracy.

This poster explores a new approach using a CIMac™ PrimaS analytical HPLC column to separate and quantify several key IVT components in a very short run time, enabling fast “at line” tracking.

The experiment involves increasing the levels of Mg2+ in the IVT reaction mix whilst using the CIMac™ PrimaS at each concentration to determine the best Mg2+ concentration range for optimum IVT kinetics.

CIMac PrimaS HPLC Column
CIMac PrimaS HPLC Column

For a copy of the poster (PDF 410 KB) CLICK HERE

Time-point analysis graph

To find out more about the CIMac™ analytical column range, or other monoliths from BIA-Separations, go to our Columns and Media page

CIMac Analytical Columns

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Posted in Chromatography