Journal: bioRxiv

Article Title: Modified self-amplifying RNA mediates robust and prolonged gene expression in the mammalian brain

doi: 10.1101/2025.10.30.685635

Figure Lengend Snippet: (A) An example brain slice showing saRNA-LNP mediated gene expression in neurons. (i) NeuN immunofluorescence from neurons (green); (ii) saRNA-LNP mediated mCherry fluorescence; (iii) GFAP immunofluorescence from astrocytes (blue); (iv) Merge, showing colocalization of mCherry and NeuN immunofluorescence (orange). (v). zoomed in view of the area highlighted by yellow box in iv. (vi). Cell profiler identification of pixels positive for NeuN (green), saRNA (red), and GFAP (blue). (B, C) Similar to A, but for example brain slices showing saRNA-LNP mediated gene expression in astrocytes. Merge showing colocalization of mCherry and GFAP immunofluorescence (purple). (D) Quantification of the percentage of saRNA-mediated mCherry expression colocalization with neurons (green) versus glia (blue) across 5 time points after saRNA injection. ***, p<0.001, Wilcoxon rank-sum test (n=10-19 fields of view per timepoint).

Article Snippet: Sections were then incubated with primary antibodies against neuronal nuclei, NeuN (rabbit, 1:500; Abcam, ab177487) and glial fibrillary acidic protein, GFAP (chicken, 1:1000; Novus Biologicals, NBP1-05198) at 4°C overnight followed by secondary antibodies Alexafluor 488 goat anti-rabbit (1:500; Invitrogen, A11008) and Alexafluor 647 goat anti-chicken (1:500; Invitrogen, A21449) for 2 hours at room temperature.

Techniques: Slice Preparation, Gene Expression, Immunofluorescence, Fluorescence, Expressing, Injection