Wyatt dynamics software

Purified cVLP:IL-1β vaccines were quality checked by Dynamic Light Scattering (DLS) and negative stain transmission electron microscopy (TEM). For DLS analysis, vaccines were spun at 16,000× g for 2 min (Eppendorf tabletop centrifuge 5424 R) before being loaded into a disposable cuvette. The samples were run with 20 acquisitions for 7 s at 25 °C using a DynaPro Nanostar (Wyatt Technologies, Santa Barbara, CA, USA). The estimated diameter of the cVLP:IL-1β particle population and the percent polydispersity (%Pd) were calculated using Wyatt DYNAMICS software (7.10.0.21) (Wyatt Technologies). For TEM, vaccines were adsorbed onto 200-mesh carbon-coated grids and stained with 2% uranyl acetate for 1 min. The excess stain was removed with filter paper. The grids were analysed using a CM 100 BioTWIN electron microscope (Philips, Amsterdam, Netherlands).

Purified PCSK9 vaccines were quality assessed by DLS and negative-stain TEM. For DLS analysis, vaccines were diluted to ~0.5 mg/mL and spun at 16,000 g for 2 min before being loaded into a disposable cuvette. The samples were run with 20 acquisitions of 7 s, at 25 °C using a DynaPro Nanostar (Wyatt Technologies, Santa Barbara, CA, USA). The estimated diameter of the cVLP-PCSK9 vaccine population and percent polydispersity (%Pd) were calculated by Wyatt DYNAMICS software (v7.10.0.21, US) (Wyatt Technologies). For TEM, vaccines were adsorbed onto 200-mesh carbon-coated grids and stained with 2% uranyl acetate. The grids were analyzed using a CM 100 BioTWIN electron microscope (Philips, Amsterdam, Netherlands), with an accelerating voltage of 80 kV.

cVLPs were mixed with a 2× molar excess of antigen, then incubated overnight at 4 °C with gentle shaking. LPS was removed from the vaccines by phase extraction using Triton X-114 as previously described [48 (link)]. Unbound antigen was removed by density gradient ultracentrifugation (OptiPrep™, Sigma-Aldrich, St. Louis, MO, USA). Fractions containing the vaccine was dialyzed using a Spectra/Por™ Cellulose Ester 1000 kDa MWCO dialysis tubing (Spectrum Chemical, New Brunswick, NJ, USA) in PBS or PBS supplemented with 0.4 M NaCl for HER2 and VAR2CSA vaccines respectively. cVLP-bound antigen concentration as well as the coupling efficiency was determined by SDS-PAGE densitometric analysis. For DLS analysis (DynaPro NanoStar, Wyatt Technology, Santa Barbara, CA, USA), the sample was spun at 16,000× g for 2 min, then loaded into a disposable cuvette. Samples were run with 20 acquisitions of 7 s each. The estimated diameter of the cVLP vaccine population and the percent polydispersity (%Pd) was calculated with Wyatt DYNAMICS software (v7.10.0.21, Wyatt Technology, Santa Barbara, CA, USA).