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golgi  (Addgene inc)
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Sacsin deletion disrupts the network of the cytolinker plectin in C6 cells. (A) Representative images of immunocytochemistry for endogenous plectin (green) in C6 cells and C6 Sacs-/- cells. (B-C) Representative immunofluorescence images showing the distribution of different organelles in C6 and C6 Sacs-/- <t>cells</t> <t>transfected</t> with <t>EYFP-Golgi7</t> (green) and immunostained with an antibody against nestin (magenta). Mitochondria were counterstained with Mitotracker (red) and nuclei with Hoechst (blue). (B) Mitochondria (red) is depleted in the juxtanuclear area in C6 Sacs-/- cells, and the <t>Golgi</t> complex (green) is in the periphery of the same region. (C) A normal compact juxtanuclear Golgi complex (green) can be seen in C6 reference cells, whereas in C6 Sacs-/- cells, it is scattered around nestin aggregates (magenta). (D) Quantification of microscopy images from 4 independent experiments (mean ± SEM). *, significant vs. C6 reference strain (p < 0.05, Student’s t-test). In 4 independent experiments, the total number of reference C6 cells counted was 588, and the total number of C6 Sacs-/- cells counted was 995. (E) Quantification of microscopy images from 3 independent experiments (mean ± SEM). The total number of transfected cells counted was 344 in the C6 reference strain and 322 in C6 Sacs-/- cells. (F) Quantification of microscopy images from 3 independent experiments (mean ± SEM). *, significant vs. C6 reference strain (p < 0.05, Student’s t-test). In 3 independent experiments, the total number of reference C6 cells counted was 464 and the total number of C6 Sacs-/- cells counted was 697. (G) Representative Western blots showing the expression patterns of the glial intermediate filament Nestin. Scale 20 μm.
Golgi, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/56590/EYFP-Golgi-7+(Plasmid+%2356590)/bio_rxiv__2024__04__15__589510-26-14-16
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Sacsin deletion disrupts the network of the cytolinker plectin in C6 cells. (A) Representative images of immunocytochemistry for endogenous plectin (green) in C6 cells and C6 Sacs-/- cells. (B-C) Representative immunofluorescence images showing the distribution of different organelles in C6 and C6 Sacs-/- cells transfected with EYFP-Golgi7 (green) and immunostained with an antibody against nestin (magenta). Mitochondria were counterstained with Mitotracker (red) and nuclei with Hoechst (blue). (B) Mitochondria (red) is depleted in the juxtanuclear area in C6 Sacs-/- cells, and the Golgi complex (green) is in the periphery of the same region. (C) A normal compact juxtanuclear Golgi complex (green) can be seen in C6 reference cells, whereas in C6 Sacs-/- cells, it is scattered around nestin aggregates (magenta). (D) Quantification of microscopy images from 4 independent experiments (mean ± SEM). *, significant vs. C6 reference strain (p < 0.05, Student’s t-test). In 4 independent experiments, the total number of reference C6 cells counted was 588, and the total number of C6 Sacs-/- cells counted was 995. (E) Quantification of microscopy images from 3 independent experiments (mean ± SEM). The total number of transfected cells counted was 344 in the C6 reference strain and 322 in C6 Sacs-/- cells. (F) Quantification of microscopy images from 3 independent experiments (mean ± SEM). *, significant vs. C6 reference strain (p < 0.05, Student’s t-test). In 3 independent experiments, the total number of reference C6 cells counted was 464 and the total number of C6 Sacs-/- cells counted was 697. (G) Representative Western blots showing the expression patterns of the glial intermediate filament Nestin. Scale 20 μm.

Journal: bioRxiv

Article Title: Subcellular, biochemical and biophysical alterations in two glial cell models of ARSACS

doi: 10.1101/2024.04.15.589510

Figure Lengend Snippet: Sacsin deletion disrupts the network of the cytolinker plectin in C6 cells. (A) Representative images of immunocytochemistry for endogenous plectin (green) in C6 cells and C6 Sacs-/- cells. (B-C) Representative immunofluorescence images showing the distribution of different organelles in C6 and C6 Sacs-/- cells transfected with EYFP-Golgi7 (green) and immunostained with an antibody against nestin (magenta). Mitochondria were counterstained with Mitotracker (red) and nuclei with Hoechst (blue). (B) Mitochondria (red) is depleted in the juxtanuclear area in C6 Sacs-/- cells, and the Golgi complex (green) is in the periphery of the same region. (C) A normal compact juxtanuclear Golgi complex (green) can be seen in C6 reference cells, whereas in C6 Sacs-/- cells, it is scattered around nestin aggregates (magenta). (D) Quantification of microscopy images from 4 independent experiments (mean ± SEM). *, significant vs. C6 reference strain (p < 0.05, Student’s t-test). In 4 independent experiments, the total number of reference C6 cells counted was 588, and the total number of C6 Sacs-/- cells counted was 995. (E) Quantification of microscopy images from 3 independent experiments (mean ± SEM). The total number of transfected cells counted was 344 in the C6 reference strain and 322 in C6 Sacs-/- cells. (F) Quantification of microscopy images from 3 independent experiments (mean ± SEM). *, significant vs. C6 reference strain (p < 0.05, Student’s t-test). In 3 independent experiments, the total number of reference C6 cells counted was 464 and the total number of C6 Sacs-/- cells counted was 697. (G) Representative Western blots showing the expression patterns of the glial intermediate filament Nestin. Scale 20 μm.

Article Snippet: When indicated, cells were transfected 24h after seeding with mammalian expression plasmids to visualize Golgi (EYFP-Golgi7, Addgene plasmids # 56590), using JetPRIME transfection reagent (Polyplus transfection, Illkirch, France) at 1:3 transfection ratios.

Techniques: Immunocytochemistry, Immunofluorescence, Transfection, Microscopy, Western Blot, Expressing