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wild-type mice c57bl/6jolahsd  (Harlan Laboratories)
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Harlan Laboratories wild-type mice c57bl/6jolahsd
Silencing of Rab35 in vivo results in altered cell polarity and liver tissue architecture. (a) Immunofluorescence images of liver tissue collected 4 d after in utero injection of Luciferase (siLuc) and Rab35 (siRab35) siRNAs formulated into LNP via vitelline vein <t>in</t> <t>E13.5</t> embryos. The square on the low-magnification images (scale bar: 500 µm) shows where the high-resolution image was taken (scale bar: 20 µm). Imaged areas are located in the liver parenchyma, devoid of bile duct cell marker Sox9. The inserts (scale bar: 20 µm) in a’ and a’’ show the difference between BC and bile duct-like lumina in LNP-siRab35 injected liver. Panel a’’’ compares tubular lumina in the parenchyma to the bile duct lumina in the portal area (Sox9-positive cells near portal veins [PV]). (b) Immunofluorescence images of liver tissues from panel a show examples of hepatocyte polarity in the control tissue (a single hepatocyte forms multiple lumina per cell) and simple apico-basal polarity in LNP-siRab35 injected liver (cells have a single apical domain oriented toward a shared lumen). (c and d) 3D reconstruction of lumina labeled with an apical marker CD13 in 100-µm-thick sections of liver tissue injected with LNP-siLuc (c) and LNP-siRab35 (d). Scale bar: 30 µm. See also . (e) Quantification of the lumen radius distribution based on the 3D reconstructions such as in c and d ( n = 3, error bars: SEM). (f) 3D reconstruction of a tubule in LNP-siRab35–injected livers shows a cylindrical lumen (green) surrounded by multiple cells. See also . (g) A cross-section of the reconstructed tubule in f in the original microscopy image shows organization of the cells around the lumen. Scale bar: 20 µm. (h) Quantification of number of cells surrounding the lumen in relation to lumen radius and position along the tubule.
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Silencing of Rab35 in vivo results in altered cell polarity and liver tissue architecture. (a) Immunofluorescence images of liver tissue collected 4 d after in utero injection of Luciferase (siLuc) and Rab35 (siRab35) siRNAs formulated into LNP via vitelline vein in E13.5 embryos. The square on the low-magnification images (scale bar: 500 µm) shows where the high-resolution image was taken (scale bar: 20 µm). Imaged areas are located in the liver parenchyma, devoid of bile duct cell marker Sox9. The inserts (scale bar: 20 µm) in a’ and a’’ show the difference between BC and bile duct-like lumina in LNP-siRab35 injected liver. Panel a’’’ compares tubular lumina in the parenchyma to the bile duct lumina in the portal area (Sox9-positive cells near portal veins [PV]). (b) Immunofluorescence images of liver tissues from panel a show examples of hepatocyte polarity in the control tissue (a single hepatocyte forms multiple lumina per cell) and simple apico-basal polarity in LNP-siRab35 injected liver (cells have a single apical domain oriented toward a shared lumen). (c and d) 3D reconstruction of lumina labeled with an apical marker CD13 in 100-µm-thick sections of liver tissue injected with LNP-siLuc (c) and LNP-siRab35 (d). Scale bar: 30 µm. See also . (e) Quantification of the lumen radius distribution based on the 3D reconstructions such as in c and d ( n = 3, error bars: SEM). (f) 3D reconstruction of a tubule in LNP-siRab35–injected livers shows a cylindrical lumen (green) surrounded by multiple cells. See also . (g) A cross-section of the reconstructed tubule in f in the original microscopy image shows organization of the cells around the lumen. Scale bar: 20 µm. (h) Quantification of number of cells surrounding the lumen in relation to lumen radius and position along the tubule.

Journal: The Journal of Cell Biology

Article Title: Anisotropic expansion of hepatocyte lumina enforced by apical bulkheads

doi: 10.1083/jcb.202103003

Figure Lengend Snippet: Silencing of Rab35 in vivo results in altered cell polarity and liver tissue architecture. (a) Immunofluorescence images of liver tissue collected 4 d after in utero injection of Luciferase (siLuc) and Rab35 (siRab35) siRNAs formulated into LNP via vitelline vein in E13.5 embryos. The square on the low-magnification images (scale bar: 500 µm) shows where the high-resolution image was taken (scale bar: 20 µm). Imaged areas are located in the liver parenchyma, devoid of bile duct cell marker Sox9. The inserts (scale bar: 20 µm) in a’ and a’’ show the difference between BC and bile duct-like lumina in LNP-siRab35 injected liver. Panel a’’’ compares tubular lumina in the parenchyma to the bile duct lumina in the portal area (Sox9-positive cells near portal veins [PV]). (b) Immunofluorescence images of liver tissues from panel a show examples of hepatocyte polarity in the control tissue (a single hepatocyte forms multiple lumina per cell) and simple apico-basal polarity in LNP-siRab35 injected liver (cells have a single apical domain oriented toward a shared lumen). (c and d) 3D reconstruction of lumina labeled with an apical marker CD13 in 100-µm-thick sections of liver tissue injected with LNP-siLuc (c) and LNP-siRab35 (d). Scale bar: 30 µm. See also . (e) Quantification of the lumen radius distribution based on the 3D reconstructions such as in c and d ( n = 3, error bars: SEM). (f) 3D reconstruction of a tubule in LNP-siRab35–injected livers shows a cylindrical lumen (green) surrounded by multiple cells. See also . (g) A cross-section of the reconstructed tubule in f in the original microscopy image shows organization of the cells around the lumen. Scale bar: 20 µm. (h) Quantification of number of cells surrounding the lumen in relation to lumen radius and position along the tubule.

Article Snippet: For primary hepatoblast isolations, embryonic livers were collected from timed-pregnant (E13.5–E14.5) wild-type mice C57BL/6JOlaHsd (Harlan Laboratories/Envigo) or C57BL6/JRj (Janvier Labs), or transgenic lines LifeAct-EGFP , ROSAmT/mG , or the in-cross of the two transgenic lines.

Techniques: In Vivo, Immunofluorescence, In Utero, Injection, Luciferase, Marker, Labeling, Microscopy