Journal
JOURNAL OF CHROMATOGRAPHY A
Volume 1487, Issue -, Pages 89-99Publisher
ELSEVIER
DOI: 10.1016/j.chroma.2016.12.085
Keywords
Vaccine; HPLC; Enveloped VLP; MALS
Funding
- European Union Seventh Framework Programme (FP7) [312004]
- Federal Ministry of Science, Research and Economy (BMWFW)
- Federal Ministry of Traffic, Innovation and Technology (bmvit)
- Styrian Business Promotion Agency SFG
- Standortagentur Tirol
- ZIT - Technology Agency of the City of Vienna through the COMET
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The rapid quantification of enveloped virus-like particles (VLPs) requires orthogonal methods to obtain reliable results. Three methods nanoparticle tracking analysis (NTA), size-exclusion HPLC (SEHPLC) with UV detection, and detection with multi-angle light scattering (MALS) for quantification of enveloped VLPs have been compared, and the lower and upper limits of detection and quantification have been evaluated. NTA directly counts the enveloped VLPs, and a particle number is obtained with a lower limit of detection (LLOD) of 1.7 x 10(7) part/mL and lower limit of quantification (LLOQ) of 3.4 x 10(8) part/mL. SE-HPLC with UV detection was calibrated with standards characterized by NTA, and a LLOD of 6.9 x 10(9) part/mL and LLOQ of 2.1 x 10(10) part/mL were found. SE-HPLC with MALS does not require a pre-calibrated sample because with a spherical model based on the Rayleigh-Gans-Debye approximation, the particle concentration can be directly deduced from the scattered light. A LLOD of 4.8 x 10(8) part/mL and LLOQ of 2.1 x 10(9) part/mL were measured and substantially lower compared to the UV method. The absolute particle concentration measured by SE-HPLC-MALS is one order of magnitude lower compared to measurement by NTA, which is explained by the wide size distribution of an enveloped VLP suspension. The model used for evaluation of light scattering data assumes monodisperse, homogeneous, and spherical particles. (C) 2017 The Authors. Published by Elsevier B.V.
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