Background: Vaccine formulations may contain visible and/or subvisible particles, which
can vary in both size and morphology. Extrinsic particles, which are particles not part of the product
such as foreign contaminants, are generally considered undesirable and should be eliminated or controlled
in injectable products. However, biological products, in particular vaccines, may also contain
particles that are inherent to the product. Here we focus on the characterization of visible and subvisible
particles in a live, replication-deficient viral vaccine candidate against HSV genital herpes in an
early developmental stage.
Method: HSV-2 viral vaccine was characterized using a panel of analytical methods, including Fourier
transform infrared spectroscopy (FTIR), sodium dodecyl sulfate-polyacrylamide gel electrophoresis
(SDS-PAGE), Western blot, liquid chromatography-mass spectrometry (LC-MS), light microscopy,
transmission electron microscopy (TEM), micro-flow imaging (MFI), dynamic light scattering (DLS),
right angle light scattering (RALS), and intrinsic fluorescence.
Results: Particles in HSV-2 vaccine typically ranged from hundreds of nanometers to hundreds of micrometers
in size and were determined to be inherent to the product. The infectious titer did not correlate
with any trend in subvisible particle concentration and size distribution as shown by DLS, MFI,
and TEM under stressed conditions. This suggested that particle changes in the submicron range were
related to HSV-2 virion structure and had direct impact on biological activity. It was also observed that
subvisible and visible particles could induce aggregation in the viral product. The temperature induced
aggregation was observed by RALS, intrinsic fluorescence, and DLS. The increase of subvisible particle
size with temperature could be fitted to a two-step thermokinetic model.
Conclusion: Visible and subvisible particles were found to be inherent to the HSV-2 viral vaccine
product. The mechanism of protein aggregation was discussed and a two-step thermokinetic aggregation
profile was proposed. The approaches reported in this study may be applied to a variety of vaccines
and other biological products, as a way to assess the consistency of the manufacturing process
and identify key product quality attributes.