title: Investigation into alternative testing methodologies for characterization of influenza virus vaccine.
abstract: The objective of this study was to explore various testing methodologies suitable for characterizing sedimented or agglomerated material. To model this, bioCSL's split influenza virus vaccine, Fluvax® was utilized. The investigation was conducted on 5 dispensed lots of commercially manufactured vaccine, formulated for the 2013 Southern Hemisphere season. Vaccine syringes were initially inspected by visual tests; the material was then aseptically pooled for characterization assessment by microscopy and several agglomeration assays. All syringes passed bioCSL's description test where any fine or large sized particles of sediment observed in the vaccine were resuspended upon shaking; inverted light microscopy verified that the sediment morphology was consistent with influenza vaccine. Electron microscopic examination of pooled vaccine material demonstrated the presence of typical influenza structures including split virus, virosomes, whole virus particles and agglomerates. An optical density turbidity assay revealed relatively high protein recoveries in the vaccine supernatant post-centrifugation treatment, thus indicative of a well-dispersed vaccine formulation. This was corroborated by particle sizing analysis using dynamic light scattering which generated reproducible volume particle size distributions of a polydisperse nature. Ultraviolet-visible absorbance profiles further confirmed the presence of some agglomerated material. Data from all methods demonstrated consistent results between all batches of vaccine. Therefore, this investigation revealed the suitability and usefulness of the various methodologies in characterizing the appearance of agglomerated vaccine material. It is suggested that such methods may be applicable and beneficial for the development of a wider spectrum of heterogeneous and agglomerated formulations to provide safe, efficacious and superior quality biopharmaceutical products.
authors.affiliation: a Influenza Research and Development; bioCSL Pty Ltd. ; Parkville , Victoria , Australia.
authors.forename: Tracy
authors.initials: T
authors.lastname: Tay
authors.forename: Catherine
authors.initials: C
authors.lastname: Agius
authors.forename: Ross
authors.initials: R
authors.lastname: Hamilton
authors.forename: Jesse
authors.initials: J
authors.lastname: Bodle
authors.forename: Steve
authors.initials: S
authors.lastname: Rockman
mesh_terms: D006801:Humans; D007252:Influenza Vaccines / Q000032:analysis / Q000737:chemistry; D008853:Microscopy; D010316:Particle Size; D013594:Syringes; D014771:Virion / Q000648:ultrastructure
chemical_list: D007252:Influenza Vaccines
keywords: AI, Agglomeration index; ANOVA, Analysis of variance; AUC, Analytical ultracentrifugation; DLS, Dynamic light scattering; EM, Electron microscopy; EP, European Pharmacopoeia; FDA, Food and Drug Administration; FFF, Field flow fractionation; HA, Hemagglutinin; ILM, Inverted light microscopy; IVV, Influenza virus vaccine; NA, Neuraminidase; OD, Optical density; ODT, Optical density turbidity; PSD, Particle size distribution; QbD, Quality by Design; RCF, Relative centrifugal force; RI, Refractive index; SEC, Size exclusion chromatography; SH, Southern Hemisphere; SLS, Static light scattering; TEM, Transmission electron microscope; TGA, Therapeutic Goods Administration; TIV, Trivalent influenza vaccine; UV, Ultraviolet; biopharmaceuticals; characterization methods; dispersed formulation; influenza vaccine; protein agglomeration; sedimentation