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Related Experiment Video

Updated: Jun 5, 2026

Highly Efficient Transfection of Primary Macrophages with In Vitro Transcribed mRNA
06:46

Highly Efficient Transfection of Primary Macrophages with In Vitro Transcribed mRNA

Published on: November 9, 2019

Continuous Purification of mRNA Produced by In Vitro Transcription Using High Performance Countercurrent Membrane

Ziqiao Wang1, Amin Javidanbardan1, Ali Behboudi1

  • 1Robert V. Waltemeyer Department of Chemical Engineering, The Pennsylvania State University, University Park, PA, USA.

Biotechnology and Bioengineering
|June 3, 2026
PubMed
Summary
This summary is machine-generated.

High-performance countercurrent membrane purification (HPCMP) effectively removes impurities like cytosine triphosphates (CTPs) and Proteinase K from messenger ribonucleic acid (mRNA) therapeutics. This continuous process shows promise for efficient mRNA purification without significant product loss or membrane fouling.

Keywords:
continuous processingmRNAmembrane purificationultrafiltration

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Area of Science:

  • Biotechnology
  • Biochemistry
  • Chemical Engineering

Background:

  • Messenger ribonucleic acid (mRNA) therapeutics require stringent purification to remove residual components from in vitro transcription.
  • Enzymes and free nucleotides are common impurities that can affect mRNA therapeutic efficacy and safety.

Purpose of the Study:

  • To evaluate the efficacy of high-performance countercurrent membrane purification (HPCMP) for purifying mRNA therapeutics.
  • To assess the removal of model impurities, cytosine triphosphates (CTPs) and Proteinase K, using HPCMP.

Main Methods:

  • HPCMP utilizing semipermeable polyethersulfone hollow fiber membranes.
  • Experiments with two model mRNA constructs and model impurities (CTPs and Proteinase K).
  • Assessment of impurity removal, mRNA yield, and process stability over time.

Main Results:

  • HPCMP achieved 99.7% removal of CTPs with no measurable mRNA loss.
  • mRNA presence reduced Proteinase K removal due to complexation, confirmed by dynamic light scattering and diafiltration.
  • Continuous operation exceeding 24 hours showed stable mRNA yield (>97%) and Proteinase K removal (>94%) without membrane fouling.

Conclusions:

  • HPCMP is a feasible method for continuous purification of mRNA therapeutics.
  • The process demonstrates high efficiency in removing nucleotide impurities and acceptable removal of enzymatic impurities.
  • The study highlights the potential of HPCMP for scalable and robust mRNA manufacturing.