LNPs were formulated as previously described4 (link),5 (link),27 (link), with the lipid components (SM-102, 1,2-distearoyl-sn-glycero-3-phosphocholine, cholesterol, and PEG-2000 at molar ratios of 50:10:38.5:1.5) being rapidly mixed with an aqueous buffer solution containing ABE8.8 or SpRY-ABE8.8 mRNA and either PAH1 gRNA or PAH2 gRNA in a 1:1 ratio by weight in 25 mM sodium acetate (pH 4.0), with an N:P ratio of 5.6. The resulting LNP formulations were subsequently dialyzed against sucrose-containing buffer, concentrated using Amicon Ultra-15 mL Centrifugal Filter Units (Millipore Sigma), sterile-filtered using 0.2-μm filters, and frozen until use. The LNPs underwent quality control for particle size (Z-Ave, hydrodynamic diameter), polydispersity index as determined by dynamic light scattering (Malvern NanoZS Zetasizer) and total RNA encapsulation as measured by the Quant-iT Ribogreen Assay (Thermo Fisher Scientific).
Amicon ultra 15 ml centrifugal filter units
Manufactured by Merck Group
The Amicon Ultra-15 mL Centrifugal Filter Units are laboratory filtration devices designed for the concentration and purification of macromolecules, such as proteins, peptides, and nucleic acids, from aqueous solutions. The units feature a regenerated cellulose membrane with a specified molecular weight cutoff, allowing the selective retention of the desired molecules while allowing smaller components to pass through during the centrifugation process.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using amicon ultra 15 ml centrifugal filter units
1
Lipid Nanoparticle Formulation and Characterization
2
Lipid Nanoparticle Formulation for mRNA Delivery
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LNPs were formulated as previously described25 (link), with the lipid components (SM-102, 1,2-distearoyl-sn-glycero-3-phosphocholine, cholesterol, and PEG-2000 at molar ratios of 50:10:38.5:1.5) being rapidly mixed with an aqueous buffer solution containing ABE8.8 mRNA and either PAH1 gRNA or PAH2 gRNA in a 1:1 ratio by weight in 25 mM sodium acetate (pH 4.0), with an N:P ratio of 5.6. The resulting LNP formulations were subsequently dialyzed against sucrose-containing buffer, concentrated using Amicon Ultra-15 mL Centrifugal Filter Units (Millipore Sigma), sterile-filtered using 0.2-µm filters, and frozen until use. The LNPs had particle sizes of 69–89 nm (Z-Ave, hydrodynamic diameter), with a polydispersity index of <0.21 as determined by dynamic light scattering (Malvern NanoZS Zetasizer) and 90–100% total RNA encapsulation as measured by the Quant-iT Ribogreen Assay (Thermo Fisher Scientific).
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