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![[Figure 3]](ju5081fig3mag.jpg)
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Figure 3
AF4 elution profiles and SAXS fractogram: system validation using model protein apoferritin [(a)–(d)] and polystyrene (PS) beads [(e)–(h)]. (a) Schematic overview depicting the various flow rate settings during the AF4 separation process of apoferritin. (b) Scattering profiles for two elution fractions shown as logI(q) versus q. Profiles are vertically shifted for clarity. The purple curve represents the scattering of the main fraction, which aligns well with the theoretical scattering profile of 24-mer apoferritin (light pink, PDB code 1IER, with CRYSOL χ2 = 1.8). (c) SAXS fractogram displaying the Integrated SAXS intensity (grey, q-range for integration 0.25–0.4 nm−1) from unsubtracted frames plotted against elution time; overlaid with UV trace (brown). Regions of interest used to generate the scattering curves in (b) are highlighted with matching colours. The inset shows an ab initio model of the main peak as spheres, overlaid with the atomistic structure of apoferritin (PDB code 1IER). (d) p(r) functions derived from the two SAXS scattering curves in (b), demonstrating the size differences between the two highlighted fractions. (e) Schematic overview depicting the separation of the PS beads containing a mixture of 20 nm and 100 nm polystyrene beads. (f) Scattering profiles obtained from the separated fractions corresponding to the 20 nm beads (magenta) and the 100 nm beads (green). (g) SAXS fractogram displaying the integrated SAXS intensity (grey, q-range for integration 0.1–0.8 nm−1) from unsubtracted frames plotted against elution time. Regions corresponding to the 20 nm and 100 nm beads are highlighted with distinct colours. (h) p(r) functions derived from the SAXS scattering curves for the 20 nm and 100 nm beads, illustrating the differences in size distribution. |
![[Figure 3]](ju5081fig3.jpg)
 | JOURNAL OF SYNCHROTRON RADIATION |
ISSN: 1600-5775
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