![]() The retention properties of ultrafiltration membranes are expressed as molecular weight cut-off (MWCO), which is based on the ability to retain > 90% of a solute of a known molecular weight (in kD). Molecules larger than the membrane pores will be retained, but not bound, at the surface of the membrane and concentrated during the ultrafiltration process. The primary basis for separation is molecular size. This fast and relatively inexpensive process can be readily executed in the laboratory by centrifugal ultrafiltration. This step exchanges the buffer and removes excess lipids and non-encapsulated mRNA to further increase the purity. This lipid mixture circumvents formulation-related challenges, allowing a quick start to setting up an mRNA-LNP laboratory. It is comprised of lipid components at defined ratios and is optimized for use with the NanoAssemblr® systems. GenVoy-Ionizable Lipid Mix (GenVoy-ILM™) is a pre-optimized Research Use Only lipid mixture designed for encapsulation and can be used at various stages of drug development from discovery to late preclinical, effectively delivering mRNA to cells with high efficiency and low toxicity.Precision Nanosystems NanoAssemblr® Ignite™ system (left) and workflow (right). The optional in-line dilution via the choice of appropriate single-use NxGen™ cartridges enhances flexibility.įig 2. Recipes are reproducible and can be stored. The setup is easy with a touch screen interface. By adjusting key parameters such as flow rate ratio, total flow rate, and dilution, lipid composition is optimized to obtain the desired LNP size and structure Formulation times are short (< 1 min/run, total flow rate up to 20 mL/min) allowing formulations to be made on-demand. NanoAssemblr® Ignite™ is the larger sibling of the Spark™ system, capable of formulating 1–20 mL. Precision Nanosystems NanoAssemblr® Spark™ system (top) and workflow (bottom). Since formulation takes barely 10 s per run, LNPs can be made on-demand for immediate use.įig 1. The petite Spark™ system can easily be placed on the bench-top or in a sterile biosafety hood. The single-use NxGen™ microfluidic mixer cartridge minimizes cross-contamination, and simple one-button operation reduces user variability. NanoAssemblr® Spark™ is the smallest system that formulates between 25–250 μL of LNP, encapsulating mRNA on the microgram scale (Fig 1). Precision Nanosystems offers a range of systems and reagents based on a microfluidic mixing technique, resulting in reproducible and high-quality LNPs.Need for a ready-to-use lipid mixture that has the right combination of lipids and lipid composition.Need for reliable and user-friendly encapsulation systems.Encapsulation efficiency can be measured using fluorescence-based, RNA-specific assays.Ĭurrent mRNA-LNP vaccines for COVID-19 must be frozen for long-term storage and may be sensitive to shaking. A high PDI indicates a broad or multimodal size distribution. Nanoparticle size and polydispersity index (PDI), which gives the size distribution of nanoparticles, can be determined by using a dynamic light scattering bioanalyzer. It’s important to consider how the combination of cationic or ionizable lipids and the lipid composition (i.e., the ratio of helper lipids and surfactants) impact mRNA encapsulation.Īfter mRNA is encapsulated, the LNPs are diluted with a suitable aqueous buffer to reduce the organic solvent content. This protective structure allows the mRNA to evade degradation by endosomes and increases stability in physiological fluid. This step facilitates self-assembly into stable nanostructures such that mRNAs are encapsulated in the interior core through interactions with ionizable lipids, cholesterol, helper lipids, and stabilizers. An aqueous buffer containing mRNAs and an organic lipid mixture are rapidly mixed in a controlled and non-turbulent manner. Microfluidic mixing is one of the most effective means to formulate lipid nanoparticles (LNPs) because of its scalability and reproducibility. Through peer-to-peer interactions, we continue to encourage the spirit of collaboration developed at a time when new modalities are being used as therapies for the first time. DYNAMIC LIGHT SCATTERING PDI HOW TOAnd they share their advice on how to scale. In this series, our scientists share their journey and insights on mRNA synthesis for research purposes. ![]()
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |