Extended Data Fig. 1: OligoD insertion into Ebola GP.
a, Size-exclusion purification and gel electrophoresis of wild-type and oligoD-modified GP. Gel was stained with Coomassie brilliant blue. b, Size-exclusion chromatography coupled with multi-angle light scattering (SEC-MALS) analysis of GP and GP-12D on a Superdex 200 column. UV and LS indicate absorbance at 280 nm and light scattering signals, respectively. c, Thermal melting temperature (Tm) of wild-type and oligoD-modified GP. Data are presented as mean ± s.d. (n = 3 samples per group). d, Alum-binding assay for wild-type or oligoD-modified GP. GP proteins were pre-mixed with alum for 1 h at room temperature and then incubated in PBS containing naïve mouse serum for 24 h at 37 °C. Upon centrifugation, the concentrations of unbound GP in the supernatant were quantified by ELISA. The rinsed pellet was analyzed by Western blotting to detect alum‐bound GP. e, Standard curve established with known concentrations of GP for reference. Data are presented as mean ± s.d. (n = 3 samples per group). f, Concentrations of unbound GP in the supernatant of GP-alum mixtures. Data are presented as mean ± s.d. (n = 3 samples per group). g, Immunogold labeling of alum complexed with GP or GP-12D. Alum alone and alum incubated with naïve mouse serum served as controls. Arrows indicate alum pellets after staining. Scale bar, 1 cm. h, Representative transmission electron micrographs of antigen-alum complexes prepared from g. Scale bar, 100 nm. i, Thermal melting profiles (left) and Tm (right) of wild-type or oligoD-modified GP in the presence of alum. Data are presented as mean ± s.d. (n = 3 samples per group). j, Detection of Adju-Phos-bound GP by Western-blot analysis. Adju-Phos was used instead of Alhydrogel in the antigen-binding assay.
