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Evaluation of Microflow Digital Imaging Particle Analysis for Sub-Visible Particles Formulated with an Opaque Vaccine
Grant E Frahm1, Alex W T Pochopsky1,2, Tessa M Clarke1,3
1Biologics and Genetic Therapies Directorate, Health Canada, Ottawa, Ontario, Canada.
Abstract:
Microflow digital imaging (MDI) has become a widely accepted method for assessing sub-visible particles in pharmaceutical formulations however, to date; no data have been presented on the utility of this methodology when formulations include opaque vaccine adjuvants. This study evaluates the ability of MDI to assess sub-visible particles under these conditions. A Fluid Imaging Technologies Inc. FlowCAM® instrument was used to assess a number of sub-visible particle types in solution with increasing concentrations of AddaVax™, a nanoscale squalene-based adjuvant. With the objective (10X) used and the limitations of the sensor resolution, the instrument was incapable of distinguishing between sub-visible particles and AddaVax™ droplets at particle sizes less than 5 μm. The instrument was capable of imaging all particle types assessed (polystyrene beads, borosilicate glass, cellulose, polyethylene protein aggregate mimics, and lysozyme protein aggregates) at sizes greater than 5 μm in concentrations of AddaVax™ up to 50% (vol:vol). Reduced edge gradients and a decrease in measured particle sizes were noted as adjuvant concentrations increased. No significant changes in particle counts were observed for polystyrene particle standards and lysozyme protein aggregates, however significant reductions in particle counts were observed for borosilicate (80% of original) and cellulose (92% of original) particles. This reduction in particle counts may be due to the opaque adjuvant masking translucent particles present in borosilicate and cellulose samples. Although the results suggest that the utility of MDI for assessing sub-visible particles in high concentrations of adjuvant may be highly dependent on particle morphology, we believe that further investigation of this methodology to assess sub-visible particles in challenging formulations is warranted.
Insights
Microflow digital imaging (MDI) can assess sub-visible particles in opaque vaccine adjuvant formulations. However, small particles (<5 μm) and some translucent particles may be obscured by opaque adjuvants like AddaVax™.
Area of Science:
- Pharmaceutical analysis
- Particle characterization
- Vaccine formulation technology
Background:
- Microflow digital imaging (MDI) is a standard for sub-visible particle analysis in pharmaceuticals.
- Limited data exists on MDI's performance with opaque vaccine adjuvants.
- Opaque adjuvants can interfere with particle detection and quantification.
Purpose of the Study:
- To evaluate the efficacy of MDI for sub-visible particle assessment in formulations containing opaque vaccine adjuvants.
- To determine the impact of increasing adjuvant concentrations on MDI's particle detection capabilities.
- To identify limitations and potential challenges of using MDI with complex vaccine formulations.
Main Methods:
- Utilized a FlowCAM® instrument for MDI analysis.
- Tested various sub-visible particle types (polystyrene beads, glass, cellulose, protein aggregates) in solutions with increasing concentrations of AddaVax™ (a nanoscale squalene-based adjuvant).
- Assessed particle imaging and counting accuracy at different adjuvant concentrations (up to 50% vol:vol).
Main Results:
- MDI could not distinguish particles <5 μm from adjuvant droplets.
- Particles >5 μm were imaged, but edge gradients and measured sizes were affected by adjuvant concentration.
- Particle counts for translucent materials (borosilicate, cellulose) significantly decreased, likely due to adjuvant masking.
- Counts for polystyrene and lysozyme aggregates remained stable.
Conclusions:
- MDI's utility for sub-visible particle analysis in high-adjuvant formulations depends on particle morphology.
- Opaque adjuvants can mask translucent particles, affecting accurate quantification.
- Further research is needed to optimize MDI for challenging vaccine formulations.
