mRNA therapeutics rely on LNP delivery vehicles, but LNP instability (aggregation, hydrolysis, etc.) hinders commercialization. LNP physicochemical integrity links to efficacy, and advanced delivery needs LNPs resistant to storage and manufacturing stress.
Our Custom Physicochemical Stability Evaluation Service accelerates mRNA development via Stability-by-Design, connecting LNP integrity to protein expression. Creative Biolabs ensures LNPs are functionally stable, providing optimal formulation/storage answers for commercial success and next-gen delivery.
Structural Integrity-Related Indicators
Component Stability-Related Indicators
Fig.1 The representative methods and schemes currently used for evaluating the stability of mRNA-LNP.1
| Detection Indicator | Detection Method | Principle & Advantage |
|---|---|---|
| Encapsulation Efficiency Change | Field-Flow Fractionation-Multi-Angle Light Scattering (FFF-MALS), Reverse-Phase High-Performance Liquid Chromatography (RP-HPLC) | FFF-MALS separates and quantifies free/encapsulated drugs; RP-HPLC calculates the proportion of leaked drugs via chromatographic peak area difference, with high sensitivity. |
| Lipid Membrane Integrity | Differential Scanning Calorimetry (DSC), Small-Angle X-Ray Scattering (SAXS) | DSC detects shifts in lipid phase transition temperature (Tm) to reflect membrane fluidity changes; SAXS analyzes lipid bilayer thickness/arrangement to visually show membrane structure damage. |
| Internal Phase Stability | Cryogenic Transmission Electron Microscopy (Cryo-TEM), X-Ray Diffraction (XRD) | Cryo-TEM directly observes internal liquid crystalline morphology; XRD confirms the retention of functional phases via characteristic diffraction peaks (e.g., d-value of HⅡ phase). |
| Lipid Degradation Products | Ultra-High Performance Liquid Chromatography-Tandem Mass Spectrometry (UPLC-MS/MS) | Separates and quantifies oxidation/hydrolysis products (e.g., lysophospholipids, lipid peroxides) with high specificity and low detection limit (ng level). |
| mRNA Integrity | Capillary Gel Electrophoresis (CGE), Reverse Transcription-Quantitative Polymerase Chain Reaction (RT-qPCR) | CGE separates intact/degraded mRNA fragments to calculate the integrity ratio; RT-qPCR reflects mRNA template activity via amplification efficiency. |
| pH Change | Precision pH Meter | Directly measures the pH of LNP suspensions; comparison with the initial pH assesses buffer system failure. |
| Required Starting Materials (2-3 Examples) | |
| 1. Full LNP Composition (Molar ratios of Ionizable, Helper, PEGylated lipids) | |
| 2. Target mRNA Sequence and desired in-house analytical data (e.g., initial DLS, EE%) | |
| 3. Target Delivery Route (e.g., IV administration, MNP/lyophilized format, or specific storage temperature) | |
| Key Steps Involved | Activities and Expected Outcomes |
|---|---|
| Stress Testing & Profile Generation | Subjecting LNP samples to tailored stress conditions (accelerated temperature, freeze-thaw cycles, mechanical shear, high pH). |
| Multi-Modal Characterization | Using DLS and high-resolution chromatography (HPLC/LC-MS) to quantify size distribution, leakage rate, and the presence of lipid degradation products (hydrolysis/oxidation). |
| Functional Validation (The Efficacy Link) | Performing robust in vitro (e.g., luciferase assay in target cells) or ex vivo protein expression assays on stressed samples. |
| Data Analysis & Optimization | Applying advanced kinetic modeling to calculate activation energy and degradation rates. |
| Final Deliverables (2-3 Examples) | |
| 1. Comprehensive Stability Report (Including all DLS and LC-MS raw and processed data) | |
| 2. Predictive Shelf-Life Model (Temperature-dependent stability curve and projected expiry) | |
| 3. Optimized Formulation Protocol (Defined lipid molar ratio and recommended storage/buffer conditions) | |
Estimated Timeframe: The typical timeframe for this comprehensive service ranges from 8 to 14 weeks, depending on the number of formulations tested and the complexity of the stress conditions required for your target delivery route.
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Our Advantage
Fully Customized Stress Testing
Design analytical pathways and tailored stress conditions (accelerated temperature, mechanical shear, freeze-thaw) to mimic your product's intended storage/delivery, including MNP fabrication or lyophilization simulation.
Integrated Multi-Modal Analytics
Seamlessly integrate DLS and high-resolution LC-MS with sensitive leakage assays to profile chemical degradation, delivering a 360-degree comprehensive analysis of LNP structural integrity.
Targeted Compositional Optimization
Provide formulation screens to quickly identify optimal molar ratios of ionizable, helper, and PEGylated lipids for maximum stability and long-term process resistance in your buffer system.
Predictive Shelf-Life Modeling
Use advanced kinetic degradation models to calculate activation energy and deliver accurate temperature-dependent shelf-life predictions, de-risking cold chain logistics and CMC submissions.
Mandatory Functional Validation
Include a final validation step linking each stability metric to sustained in vitro/ex vivo protein expression, ensuring stable LNPs remain potent therapeutics.
CMC-Ready Documentation
Generate all data, protocols, and analyses under a robust quality system, providing QbD documentation for successful regulatory approval.
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A: DLS and basic EE% are necessary but insufficient—they miss subtle changes (e.g., low-level fusion, chemical degradation) not affecting average particle size. Our multi-modal approach (DLS + LC-MS + Functional Assay) catches invisible issues (cargo leakage, lipid breakdown) that make formulations functionally inert, providing data critical for regulatory confidence.
A: Yes. We offer Process-Induced Stress testing (simulating lyophilization/MNP fabrication) beyond standard cold storage. We identify optimal ionizable lipid molar ratios and stabilizing excipients to keep LNPs structurally sound and potent post-reconstitution.
A: The most critical factor is chemical degradation rate (hydrolysis/oxidation) via LC-MS. This enables Arrhenius kinetics to build accurate temperature-dependent shelf-life models, justifying refrigerated over ultra-cold storage.
A: No. It's also used to troubleshoot unstable existing formulations (e.g., lab-to-GMP scale-up). We pinpoint failure mechanisms (e.g., shear-induced aggregation, buffer pH-related leakage) and provide corrective action plans.
A: Our Biological Validation uses target-relevant cell lines/ex vivo models. We collaborate with you to select appropriate assays and endpoints, ensuring that stability data reflects the LNP's actual therapeutic performance.
Creative Biolabs offers an integrated solution to the LNP stability crisis, focusing on a Stability-by-Design strategy that optimizes lipid composition to resist storage and manufacturing stresses. Our service guarantees that your mRNA therapeutic maintains both physicochemical integrity and functional potency, accelerating your path to commercial viability.
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