Figure 1
From: Reversible off and on switching of prion infectivity via removing and reinstalling prion sialylation
Experimental design and 2D analysis of sialylation status.
(A) Experimental design illustrating generation of desialylated and re-sialylated SSLOW material. To produce desialylated SSLOW, serial dsPMCAb reactions were seeded with brain-derived SSLOW and conducted using sialidase-treated NBH to a final dilution of SSLOW brain material of 10−24-fold. To produce re-sialylated SSLOW, serial rsPMCAb was seeded with dsPMCAb products and conducted using non-treated NBH to a final dilution of original SSLOW brain material 10−34-fold. To produce reference PMCAb-derived material, serial PMCAb reaction was seeded with brain-derived SSLOW and conducted using non-treated NBH to a final dilution of SSLOW brain material 10−27-fold. Animals were inoculated IC with 10-fold diluted PMCAb, dsPMCAb- or rsPMCAb-derived material. (B) 2D Western blot analysis of SSLOW brain-, PMCAb-, dsPMCAb- and rsPMCAb-derived material. Black and white triangles mark diglycosylated and monoglycosylated glycoforms, respectively, whereas arrows mark the unglycosylated form. All blots were stained with 3F4 antibody. (C) Sialylation profiles of diglycosylated isoforms of SSLOW brain- (solid thin line), PMCAb- (solid bold line), dsPMCAb- (dashed line) and rsPMCAb-derived material (dotted line). Profiles were built as described in Materials and Methods using results of 2D Western blots.
