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  • 99
    Vector Laboratories dapi
    Dapi, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 99/100, based on 91325 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dapi/product/Vector Laboratories
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    99
    Thermo Fisher dapi
    Cohort of cells born at E11.5. A-E: Coronal sections of E12.5 (A), 13.0 (B) and 15.5 (C) brains labeled at E11.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex of E12.5 through E15.5 were shown in (D) and (E), respectively. See also Figure 3S for low magnification pictures of brains fixed at E12.5 through E15.5. At E12.0, most of the labeled cells were located in the VZ, and some cells were in the CSPG-positive PP in both the dorsomedial and dorsolateral cortex (D, E, arrowheads, Figure S3A). At E12.5 and 13.0, more labeled cells were found in the PP in both dorsomedial (A, B, D, Figures S3B, S3C) and dorsolateral cortex (A, B, E, Figures S3B, S3C). At E 13.5, in the dorsomedial cortex, where PP splitting does not occur yet at this stage, many neurons reached the PP just beneath the meninges (D, Figure S3D). Many labeled cells were located in the newly formed CP and intermediate zone (IZ) in the dorsolateral cortex (E, Figure S3D). At E14.5, many cells were in the newly formed CP in both dorsomedial and dorsolateral cortex (D, E, Figure S3E). At E15.5, many cells were in the lower part of CP and, to lesser extent, MZ (C, D, E, Figures S3F, S3G, S3H). Some cells were also found in the SP in the dorsolateral cortex (C, E, Figures S3F, S3H). <t>DAPI,</t> <t>4’,6-diamidino-2-phenylindole;</t> FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).
    Dapi, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 156002 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dapi/product/Thermo Fisher
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    dapi - by Bioz Stars, 2020-10
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    99
    Millipore dapi
    Cohort of cells born at E11.5. A-E: Coronal sections of E12.5 (A), 13.0 (B) and 15.5 (C) brains labeled at E11.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex of E12.5 through E15.5 were shown in (D) and (E), respectively. See also Figure 3S for low magnification pictures of brains fixed at E12.5 through E15.5. At E12.0, most of the labeled cells were located in the VZ, and some cells were in the CSPG-positive PP in both the dorsomedial and dorsolateral cortex (D, E, arrowheads, Figure S3A). At E12.5 and 13.0, more labeled cells were found in the PP in both dorsomedial (A, B, D, Figures S3B, S3C) and dorsolateral cortex (A, B, E, Figures S3B, S3C). At E 13.5, in the dorsomedial cortex, where PP splitting does not occur yet at this stage, many neurons reached the PP just beneath the meninges (D, Figure S3D). Many labeled cells were located in the newly formed CP and intermediate zone (IZ) in the dorsolateral cortex (E, Figure S3D). At E14.5, many cells were in the newly formed CP in both dorsomedial and dorsolateral cortex (D, E, Figure S3E). At E15.5, many cells were in the lower part of CP and, to lesser extent, MZ (C, D, E, Figures S3F, S3G, S3H). Some cells were also found in the SP in the dorsolateral cortex (C, E, Figures S3F, S3H). <t>DAPI,</t> <t>4’,6-diamidino-2-phenylindole;</t> FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).
    Dapi, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 87206 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dapi/product/Millipore
    Average 99 stars, based on 87206 article reviews
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    dapi - by Bioz Stars, 2020-10
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    99
    Thermo Fisher 4 6 diamidino 2 phenylindole dapi
    Cohort of cells born at E11.5. A-E: Coronal sections of E12.5 (A), 13.0 (B) and 15.5 (C) brains labeled at E11.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex of E12.5 through E15.5 were shown in (D) and (E), respectively. See also Figure 3S for low magnification pictures of brains fixed at E12.5 through E15.5. At E12.0, most of the labeled cells were located in the VZ, and some cells were in the CSPG-positive PP in both the dorsomedial and dorsolateral cortex (D, E, arrowheads, Figure S3A). At E12.5 and 13.0, more labeled cells were found in the PP in both dorsomedial (A, B, D, Figures S3B, S3C) and dorsolateral cortex (A, B, E, Figures S3B, S3C). At E 13.5, in the dorsomedial cortex, where PP splitting does not occur yet at this stage, many neurons reached the PP just beneath the meninges (D, Figure S3D). Many labeled cells were located in the newly formed CP and intermediate zone (IZ) in the dorsolateral cortex (E, Figure S3D). At E14.5, many cells were in the newly formed CP in both dorsomedial and dorsolateral cortex (D, E, Figure S3E). At E15.5, many cells were in the lower part of CP and, to lesser extent, MZ (C, D, E, Figures S3F, S3G, S3H). Some cells were also found in the SP in the dorsolateral cortex (C, E, Figures S3F, S3H). <t>DAPI,</t> <t>4’,6-diamidino-2-phenylindole;</t> FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).
    4 6 Diamidino 2 Phenylindole Dapi, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 16080 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/4 6 diamidino 2 phenylindole dapi/product/Thermo Fisher
    Average 99 stars, based on 16080 article reviews
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    4 6 diamidino 2 phenylindole dapi - by Bioz Stars, 2020-10
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    99
    Millipore 4 6 diamidino 2 phenylindole dapi
    Cohort of cells born at E11.5. A-E: Coronal sections of E12.5 (A), 13.0 (B) and 15.5 (C) brains labeled at E11.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex of E12.5 through E15.5 were shown in (D) and (E), respectively. See also Figure 3S for low magnification pictures of brains fixed at E12.5 through E15.5. At E12.0, most of the labeled cells were located in the VZ, and some cells were in the CSPG-positive PP in both the dorsomedial and dorsolateral cortex (D, E, arrowheads, Figure S3A). At E12.5 and 13.0, more labeled cells were found in the PP in both dorsomedial (A, B, D, Figures S3B, S3C) and dorsolateral cortex (A, B, E, Figures S3B, S3C). At E 13.5, in the dorsomedial cortex, where PP splitting does not occur yet at this stage, many neurons reached the PP just beneath the meninges (D, Figure S3D). Many labeled cells were located in the newly formed CP and intermediate zone (IZ) in the dorsolateral cortex (E, Figure S3D). At E14.5, many cells were in the newly formed CP in both dorsomedial and dorsolateral cortex (D, E, Figure S3E). At E15.5, many cells were in the lower part of CP and, to lesser extent, MZ (C, D, E, Figures S3F, S3G, S3H). Some cells were also found in the SP in the dorsolateral cortex (C, E, Figures S3F, S3H). <t>DAPI,</t> <t>4’,6-diamidino-2-phenylindole;</t> FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).
    4 6 Diamidino 2 Phenylindole Dapi, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 12466 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Beyotime dapi
    Akt2 promotes NKT17 lineage differentiation. Cells were from WT and Akt2 KO mice. (A) Intracellular staining of PLZF, RORγt, T-bet, and GATA3 in iNKT cells from the thymus. Percentages and absolute numbers of NKT1, NKT2, NKT17 cells in the thymus ( n = 6 mice per group) (B) and spleen ( n = 4 mice per group) (C) from WT, KO, WT chimera mice ( n = 3) and Akt2 KO chimera mice ( n = 3). (D–F) Cytokine production by iNKT cells (gated on CD1d-PBS57 + TCRβ + cells) from the thymus ( n = 5 mice per group) and spleen ( n = 5 mice per group) after stimulation with PMA plus ionomycin for 5 h. (G,H) Cytokine production by iNKT cells from WT and Akt2 KO thymocytes stimulated with <t>α-GalCer</t> for 72 h and with PMA plus ionomycin in the last 5 h ( n = 4 mice per group). (I) Critical role of Akt2 for PLZF localization to the nuclear bodies. MACS—enriched and FACS—sorted NKT17 cells from WT and Akt2 KO thymocytes( n = 3) were fixed and stained with a mouse anti-PLZF and Actin as primary antibody, detected with an Goat anti mouse secondary antibody. The nuclei were stained with <t>DAPI.</t> (J) Mean fluorescent intensity of cytoplasm and nucleus. * p
    Dapi, supplied by Beyotime, used in various techniques. Bioz Stars score: 94/100, based on 7252 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Vector Laboratories vectashield mounting medium
    Akt2 promotes NKT17 lineage differentiation. Cells were from WT and Akt2 KO mice. (A) Intracellular staining of PLZF, RORγt, T-bet, and GATA3 in iNKT cells from the thymus. Percentages and absolute numbers of NKT1, NKT2, NKT17 cells in the thymus ( n = 6 mice per group) (B) and spleen ( n = 4 mice per group) (C) from WT, KO, WT chimera mice ( n = 3) and Akt2 KO chimera mice ( n = 3). (D–F) Cytokine production by iNKT cells (gated on CD1d-PBS57 + TCRβ + cells) from the thymus ( n = 5 mice per group) and spleen ( n = 5 mice per group) after stimulation with PMA plus ionomycin for 5 h. (G,H) Cytokine production by iNKT cells from WT and Akt2 KO thymocytes stimulated with <t>α-GalCer</t> for 72 h and with PMA plus ionomycin in the last 5 h ( n = 4 mice per group). (I) Critical role of Akt2 for PLZF localization to the nuclear bodies. MACS—enriched and FACS—sorted NKT17 cells from WT and Akt2 KO thymocytes( n = 3) were fixed and stained with a mouse anti-PLZF and Actin as primary antibody, detected with an Goat anti mouse secondary antibody. The nuclei were stained with <t>DAPI.</t> (J) Mean fluorescent intensity of cytoplasm and nucleus. * p
    Vectashield Mounting Medium, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 99/100, based on 63344 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/vectashield mounting medium/product/Vector Laboratories
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    vectashield mounting medium - by Bioz Stars, 2020-10
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    99
    SouthernBiotech dapi fluoromount g
    Akt2 promotes NKT17 lineage differentiation. Cells were from WT and Akt2 KO mice. (A) Intracellular staining of PLZF, RORγt, T-bet, and GATA3 in iNKT cells from the thymus. Percentages and absolute numbers of NKT1, NKT2, NKT17 cells in the thymus ( n = 6 mice per group) (B) and spleen ( n = 4 mice per group) (C) from WT, KO, WT chimera mice ( n = 3) and Akt2 KO chimera mice ( n = 3). (D–F) Cytokine production by iNKT cells (gated on CD1d-PBS57 + TCRβ + cells) from the thymus ( n = 5 mice per group) and spleen ( n = 5 mice per group) after stimulation with PMA plus ionomycin for 5 h. (G,H) Cytokine production by iNKT cells from WT and Akt2 KO thymocytes stimulated with <t>α-GalCer</t> for 72 h and with PMA plus ionomycin in the last 5 h ( n = 4 mice per group). (I) Critical role of Akt2 for PLZF localization to the nuclear bodies. MACS—enriched and FACS—sorted NKT17 cells from WT and Akt2 KO thymocytes( n = 3) were fixed and stained with a mouse anti-PLZF and Actin as primary antibody, detected with an Goat anti mouse secondary antibody. The nuclei were stained with <t>DAPI.</t> (J) Mean fluorescent intensity of cytoplasm and nucleus. * p
    Dapi Fluoromount G, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 99/100, based on 3530 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    dapi fluoromount g - by Bioz Stars, 2020-10
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    99
    Thermo Fisher prolong gold antifade mountant
    Akt2 promotes NKT17 lineage differentiation. Cells were from WT and Akt2 KO mice. (A) Intracellular staining of PLZF, RORγt, T-bet, and GATA3 in iNKT cells from the thymus. Percentages and absolute numbers of NKT1, NKT2, NKT17 cells in the thymus ( n = 6 mice per group) (B) and spleen ( n = 4 mice per group) (C) from WT, KO, WT chimera mice ( n = 3) and Akt2 KO chimera mice ( n = 3). (D–F) Cytokine production by iNKT cells (gated on CD1d-PBS57 + TCRβ + cells) from the thymus ( n = 5 mice per group) and spleen ( n = 5 mice per group) after stimulation with PMA plus ionomycin for 5 h. (G,H) Cytokine production by iNKT cells from WT and Akt2 KO thymocytes stimulated with <t>α-GalCer</t> for 72 h and with PMA plus ionomycin in the last 5 h ( n = 4 mice per group). (I) Critical role of Akt2 for PLZF localization to the nuclear bodies. MACS—enriched and FACS—sorted NKT17 cells from WT and Akt2 KO thymocytes( n = 3) were fixed and stained with a mouse anti-PLZF and Actin as primary antibody, detected with an Goat anti mouse secondary antibody. The nuclei were stained with <t>DAPI.</t> (J) Mean fluorescent intensity of cytoplasm and nucleus. * p
    Prolong Gold Antifade Mountant, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 17297 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    dapi  (Abcam)
    99
    Abcam dapi
    Akt2 promotes NKT17 lineage differentiation. Cells were from WT and Akt2 KO mice. (A) Intracellular staining of PLZF, RORγt, T-bet, and GATA3 in iNKT cells from the thymus. Percentages and absolute numbers of NKT1, NKT2, NKT17 cells in the thymus ( n = 6 mice per group) (B) and spleen ( n = 4 mice per group) (C) from WT, KO, WT chimera mice ( n = 3) and Akt2 KO chimera mice ( n = 3). (D–F) Cytokine production by iNKT cells (gated on CD1d-PBS57 + TCRβ + cells) from the thymus ( n = 5 mice per group) and spleen ( n = 5 mice per group) after stimulation with PMA plus ionomycin for 5 h. (G,H) Cytokine production by iNKT cells from WT and Akt2 KO thymocytes stimulated with <t>α-GalCer</t> for 72 h and with PMA plus ionomycin in the last 5 h ( n = 4 mice per group). (I) Critical role of Akt2 for PLZF localization to the nuclear bodies. MACS—enriched and FACS—sorted NKT17 cells from WT and Akt2 KO thymocytes( n = 3) were fixed and stained with a mouse anti-PLZF and Actin as primary antibody, detected with an Goat anti mouse secondary antibody. The nuclei were stained with <t>DAPI.</t> (J) Mean fluorescent intensity of cytoplasm and nucleus. * p
    Dapi, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 3233 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Santa Cruz Biotechnology dapi
    Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for the time courses of 20 hours (A–D), 30 hours (E–H), and 10 days (I–L). The expressions of GFAP, βTubulin, Nestin, and c-Fos were detected in differentiated (B, F, J, D, H, L) and undifferentiated (A, E, I, C, G, K) TH + rBM-MSCs. Nuclei were shown by <t>DAPI</t> in blue staining. Scale bar: 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, <t>4´,6-diamidino-2-phenylindole.</t>
    Dapi, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 94/100, based on 3226 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore 4 6 diamidino 2 phenylindole dihydrochloride
    Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for the time courses of 20 hours (A–D), 30 hours (E–H), and 10 days (I–L). The expressions of GFAP, βTubulin, Nestin, and c-Fos were detected in differentiated (B, F, J, D, H, L) and undifferentiated (A, E, I, C, G, K) TH + rBM-MSCs. Nuclei were shown by <t>DAPI</t> in blue staining. Scale bar: 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, <t>4´,6-diamidino-2-phenylindole.</t>
    4 6 Diamidino 2 Phenylindole Dihydrochloride, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 3248 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher dapi 4 6 diamidino 2 phenylindole dihydrochloride
    Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for the time courses of 20 hours (A–D), 30 hours (E–H), and 10 days (I–L). The expressions of GFAP, βTubulin, Nestin, and c-Fos were detected in differentiated (B, F, J, D, H, L) and undifferentiated (A, E, I, C, G, K) TH + rBM-MSCs. Nuclei were shown by <t>DAPI</t> in blue staining. Scale bar: 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, <t>4´,6-diamidino-2-phenylindole.</t>
    Dapi 4 6 Diamidino 2 Phenylindole Dihydrochloride, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1614 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Cell Signaling Technology Inc 4 6 diamidino 2 phenylindole
    Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for the time courses of 20 hours (A–D), 30 hours (E–H), and 10 days (I–L). The expressions of GFAP, βTubulin, Nestin, and c-Fos were detected in differentiated (B, F, J, D, H, L) and undifferentiated (A, E, I, C, G, K) TH + rBM-MSCs. Nuclei were shown by <t>DAPI</t> in blue staining. Scale bar: 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, <t>4´,6-diamidino-2-phenylindole.</t>
    4 6 Diamidino 2 Phenylindole, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 261 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Carl Zeiss 4 6 diamidino 2 phenylindole dapi
    The unfolded protein response (UPR) is an early event during in vitro hepatic stellate cell (HSC) activation. a Bright-field images of freshly isolated mouse HSCs (mHSCs), plated on plastic culture dishes, were taken after 2, 10 and 24 h and 4, 7 and 10 days in culture (upper right panel). At the same time points, expression of HSC activation markers Acta2 and Lox was determined at the mRNA level by quantitative real-time polymerase chain reaction (qPCR; upper left panel). The lower panel shows mRNA levels of UPR genes Bip , Chop , Xbp1s , Atf4 and Herpud1 at the indicated time points during HSC culture; n = 3. b Immunofluorescent staining for Bip (green) of HSCs fixed at different time points during cultivation. <t>4′,6-Diamidino-2-phenylindole</t> <t>(DAPI;</t> blue) was used as a nuclear staining. Graph representing the quantification of % mean intensity in at least 25 cells for each time point. c Western blot analysis of HSC activation markers PDGFRβ and α-smooth muscle actin (α-SMA) and of UPR marker BIP. GAPDH was used as a loading control. Cells were collected at the indicated time points, and samples for the 0 h time point were not seeded, but lysed immediately after cell isolation. Images and graphics are representatives of at least two different HSC isolations. * P
    4 6 Diamidino 2 Phenylindole Dapi, supplied by Carl Zeiss, used in various techniques. Bioz Stars score: 94/100, based on 2994 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Dojindo Labs dapi
    High-power-magnification optical microscopy images of a 4-year-old male monkey showing <t>immunostaining</t> of the fovea for CD 117 (red) and CD44 (green) in the foveola (vertical cross section). ( A ) Immunostaining for CD117 is found predominantly in the interphotoreceptor matrix (white arrowheads). ( B ) Immunostaining for CD44 is found predominantly in the Müller cell apical microvilli in the fovea (unfilled arrowheads) except the foveolar region (white arrow) and in the interphotoreceptor matrix (white arrowheads). ( C ) <t>DAPI</t> staining (blue). ( D ) In the double staining for CD117 and CD44, colocalization yellowish regions, is visible in the interphotoreceptor matrix of the fovea (white arrowheads). Discontinuity of Müller cell apical microvilli is observed in the foveolar region the foveolar region (white arrow).
    Dapi, supplied by Dojindo Labs, used in various techniques. Bioz Stars score: 93/100, based on 1436 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Agilent technologies dapi
    Coexpression of p63 with KRT5 and KRT14 in the IPF lung. a , b Sections from IPF human lungs (5 patients) were co-stained either for KRT5 ( green ) and p63 ( red ) ( a left ; b left ), or KRT14 ( red ) and p63 ( green ) ( a right ; b right ) and <t>counterstained</t> with <t>DAPI</t> ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining, for the conducting ( a ) and distal ( b ) healthy airways. c Characteristic KRT5 + pod, stained for KRT5 ( green ) and p63 (red). Arrow: KRT5 + p63 - cell. Full bars = 500 μm. Scattered bars = 100 μm
    Dapi, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 92/100, based on 4802 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore 6 diamidino 2 phenylindole dapi
    Coexpression of p63 with KRT5 and KRT14 in the IPF lung. a , b Sections from IPF human lungs (5 patients) were co-stained either for KRT5 ( green ) and p63 ( red ) ( a left ; b left ), or KRT14 ( red ) and p63 ( green ) ( a right ; b right ) and <t>counterstained</t> with <t>DAPI</t> ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining, for the conducting ( a ) and distal ( b ) healthy airways. c Characteristic KRT5 + pod, stained for KRT5 ( green ) and p63 (red). Arrow: KRT5 + p63 - cell. Full bars = 500 μm. Scattered bars = 100 μm
    6 Diamidino 2 Phenylindole Dapi, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1640 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cohort of cells born at E11.5. A-E: Coronal sections of E12.5 (A), 13.0 (B) and 15.5 (C) brains labeled at E11.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex of E12.5 through E15.5 were shown in (D) and (E), respectively. See also Figure 3S for low magnification pictures of brains fixed at E12.5 through E15.5. At E12.0, most of the labeled cells were located in the VZ, and some cells were in the CSPG-positive PP in both the dorsomedial and dorsolateral cortex (D, E, arrowheads, Figure S3A). At E12.5 and 13.0, more labeled cells were found in the PP in both dorsomedial (A, B, D, Figures S3B, S3C) and dorsolateral cortex (A, B, E, Figures S3B, S3C). At E 13.5, in the dorsomedial cortex, where PP splitting does not occur yet at this stage, many neurons reached the PP just beneath the meninges (D, Figure S3D). Many labeled cells were located in the newly formed CP and intermediate zone (IZ) in the dorsolateral cortex (E, Figure S3D). At E14.5, many cells were in the newly formed CP in both dorsomedial and dorsolateral cortex (D, E, Figure S3E). At E15.5, many cells were in the lower part of CP and, to lesser extent, MZ (C, D, E, Figures S3F, S3G, S3H). Some cells were also found in the SP in the dorsolateral cortex (C, E, Figures S3F, S3H). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Cohort of cells born at E11.5. A-E: Coronal sections of E12.5 (A), 13.0 (B) and 15.5 (C) brains labeled at E11.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex of E12.5 through E15.5 were shown in (D) and (E), respectively. See also Figure 3S for low magnification pictures of brains fixed at E12.5 through E15.5. At E12.0, most of the labeled cells were located in the VZ, and some cells were in the CSPG-positive PP in both the dorsomedial and dorsolateral cortex (D, E, arrowheads, Figure S3A). At E12.5 and 13.0, more labeled cells were found in the PP in both dorsomedial (A, B, D, Figures S3B, S3C) and dorsolateral cortex (A, B, E, Figures S3B, S3C). At E 13.5, in the dorsomedial cortex, where PP splitting does not occur yet at this stage, many neurons reached the PP just beneath the meninges (D, Figure S3D). Many labeled cells were located in the newly formed CP and intermediate zone (IZ) in the dorsolateral cortex (E, Figure S3D). At E14.5, many cells were in the newly formed CP in both dorsomedial and dorsolateral cortex (D, E, Figure S3E). At E15.5, many cells were in the lower part of CP and, to lesser extent, MZ (C, D, E, Figures S3F, S3G, S3H). Some cells were also found in the SP in the dorsolateral cortex (C, E, Figures S3F, S3H). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling

    Cohort of cells born at E12.5. A-H: Coronal sections of E13.0 (A), 13.5 (B), 14.0 (C), 14.5 (D), 15.5 (E), and 16.5 (F) brains labeled at E12.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex were shown in (G) and (H), respectively. In the dorsomedial cortex at E13.0, many labeled cells were in the VZ, but a small number of labeled cells were also found in the PP (A, G, Figure S4G). At E13.5, more labeled cells were in the PP in addition to the VZ in the dorsomedial cortex (B, G). At this stage, the incipient CP appears in the dorsolateral cortex, and many labeled neurons were migrating in the IZ (B, H). In the dorsomedial cortex of E14.0, when the incipient CP is beginning to be formed, some labeled cells reached just beneath the meningeal surface, while others seemed to be still migrating (C, G). In the dorsolateral cortex, too, many neurons reached the superficial part of the CP, while others were still migrating in the IZ and CP (C, H). At E14.5, labeled cells in the dorsomedial cortex began to be oriented radially just beneath the MZ (D, G). In the dorsolateral cortex, many strongly labeled cells were located in the cortical plate in addition to the IZ (D, H). At E 15.5, most of the labeled cells distributed not only the superficial CP, but also in the deep part of the CP in both the dorsomedial and dorsolateral CP, suggesting that some of them began to move deeper (E, G, H). At E16.5, the main population of the labeled cells was located in the somewhat deeper part of the CP in both the dorsomedial and dorsolateral cortex. In the dorsomedial cortex, many labeled cells were also distributed in the SP. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-F), 50 μm (G, H).

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Cohort of cells born at E12.5. A-H: Coronal sections of E13.0 (A), 13.5 (B), 14.0 (C), 14.5 (D), 15.5 (E), and 16.5 (F) brains labeled at E12.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex were shown in (G) and (H), respectively. In the dorsomedial cortex at E13.0, many labeled cells were in the VZ, but a small number of labeled cells were also found in the PP (A, G, Figure S4G). At E13.5, more labeled cells were in the PP in addition to the VZ in the dorsomedial cortex (B, G). At this stage, the incipient CP appears in the dorsolateral cortex, and many labeled neurons were migrating in the IZ (B, H). In the dorsomedial cortex of E14.0, when the incipient CP is beginning to be formed, some labeled cells reached just beneath the meningeal surface, while others seemed to be still migrating (C, G). In the dorsolateral cortex, too, many neurons reached the superficial part of the CP, while others were still migrating in the IZ and CP (C, H). At E14.5, labeled cells in the dorsomedial cortex began to be oriented radially just beneath the MZ (D, G). In the dorsolateral cortex, many strongly labeled cells were located in the cortical plate in addition to the IZ (D, H). At E 15.5, most of the labeled cells distributed not only the superficial CP, but also in the deep part of the CP in both the dorsomedial and dorsolateral CP, suggesting that some of them began to move deeper (E, G, H). At E16.5, the main population of the labeled cells was located in the somewhat deeper part of the CP in both the dorsomedial and dorsolateral cortex. In the dorsomedial cortex, many labeled cells were also distributed in the SP. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-F), 50 μm (G, H).

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling

    Regional differences in neuronal migration in the cerebral cortex revealed by FT. A-C: To visualize migration profile of the whole telencephalon, CFSE was injected into the ventricle of the E14.5 embryos and 5-Ethynyl-2’-deoxyuridine (EdU) was injected into the peritoneal cavity of the mother at the end of the surgery. Harvested at E16.5, many cells labeled with FT reached the superficial part of the CP in the dorsomedial cortex (cyan dotted line), while almost no cells reached the CP in the dorsolateral cortex (A, C). In the dorsolateral cortex, many neurons were just below the subplate (SP) (yellow dotted line). Such a clear difference in neuronal migration was not detected by EdU (B, C). D: FT labeling was performed at E14.0 and slice culture was prepared at E14.5. Labeled cells left the VZ and migrate in the MAZ in multipolar morphology (10:08-25:21). They gradually obtained polarity and migrate in the intermediate zone (20:17-30:25) and reached just below the SP (relatively dark band in the transmitted light channel, highlighted by white arrows). Neurons in the dorsomedial cortex (more medial than the magenta arrow) migrate smoothly to reach the most superficial part of the cortical plate(25:21-30:25), while in the dorsolateral cortex (more lateral than the magenta arrow), neurons seemed to sojourn transiently below the SP (clear in the regions lateral than the magenta arrow)(30:25-35:29). These cells subsequently migrated into the CP in the locomotion mode (35:29-40:34). E: FAST 3D imaging of E16.5 brains in which FT labeling was performed at E14.5. Anterior and posterior representative sections were shown in addition to a section at the interventricular foramen. Supplemental Movie 1 shows a whole 3D movie taken from this brain. EdU, 5-Ethynyl-2’-deoxyuridine; DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; IP, intraperitoneal injection; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; M, medial; L, lateral; D, dorsal; V, ventral. Scale bars, 200 μm.

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Regional differences in neuronal migration in the cerebral cortex revealed by FT. A-C: To visualize migration profile of the whole telencephalon, CFSE was injected into the ventricle of the E14.5 embryos and 5-Ethynyl-2’-deoxyuridine (EdU) was injected into the peritoneal cavity of the mother at the end of the surgery. Harvested at E16.5, many cells labeled with FT reached the superficial part of the CP in the dorsomedial cortex (cyan dotted line), while almost no cells reached the CP in the dorsolateral cortex (A, C). In the dorsolateral cortex, many neurons were just below the subplate (SP) (yellow dotted line). Such a clear difference in neuronal migration was not detected by EdU (B, C). D: FT labeling was performed at E14.0 and slice culture was prepared at E14.5. Labeled cells left the VZ and migrate in the MAZ in multipolar morphology (10:08-25:21). They gradually obtained polarity and migrate in the intermediate zone (20:17-30:25) and reached just below the SP (relatively dark band in the transmitted light channel, highlighted by white arrows). Neurons in the dorsomedial cortex (more medial than the magenta arrow) migrate smoothly to reach the most superficial part of the cortical plate(25:21-30:25), while in the dorsolateral cortex (more lateral than the magenta arrow), neurons seemed to sojourn transiently below the SP (clear in the regions lateral than the magenta arrow)(30:25-35:29). These cells subsequently migrated into the CP in the locomotion mode (35:29-40:34). E: FAST 3D imaging of E16.5 brains in which FT labeling was performed at E14.5. Anterior and posterior representative sections were shown in addition to a section at the interventricular foramen. Supplemental Movie 1 shows a whole 3D movie taken from this brain. EdU, 5-Ethynyl-2’-deoxyuridine; DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; IP, intraperitoneal injection; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; M, medial; L, lateral; D, dorsal; V, ventral. Scale bars, 200 μm.

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Migration, Injection, Labeling, Imaging

    Regional differences in neuronal migration in reeler mutants and Gbx2 -/- mice. A-B: In wildtype brains, Nurr1+ cells were observed in the SP (A), while in reeler mice, Nurr1+ cells were mostly observed in the superplate, or beneath the meninges (B). In contrast to wildtype mice clearly showing regional differences in neuronal migration (A), regional differences were not clear in reeler mice (B). FT was performed at E14.5 and fixed at E16.5. C-D: In Gbx2 +/- brain, Netrin G1-positive thalamocortical axons run the SP (C1). In Gbx2 -/- brain, Netrin G1-positive thalamocortical axons were almost absent in the cortex (C2). In both cases, many neurons were observed just beneath the SP. FT was performed at E14.5 and fixed 50 hour later. Coronal sections slightly caudal to the main part of the interventricular foramina were shown to evaluate the thalamus at the same sections. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; M, medial; L, lateral; D, dorsal; V, ventral. Scale bars, 200 μm.

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Regional differences in neuronal migration in reeler mutants and Gbx2 -/- mice. A-B: In wildtype brains, Nurr1+ cells were observed in the SP (A), while in reeler mice, Nurr1+ cells were mostly observed in the superplate, or beneath the meninges (B). In contrast to wildtype mice clearly showing regional differences in neuronal migration (A), regional differences were not clear in reeler mice (B). FT was performed at E14.5 and fixed at E16.5. C-D: In Gbx2 +/- brain, Netrin G1-positive thalamocortical axons run the SP (C1). In Gbx2 -/- brain, Netrin G1-positive thalamocortical axons were almost absent in the cortex (C2). In both cases, many neurons were observed just beneath the SP. FT was performed at E14.5 and fixed 50 hour later. Coronal sections slightly caudal to the main part of the interventricular foramina were shown to evaluate the thalamus at the same sections. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; M, medial; L, lateral; D, dorsal; V, ventral. Scale bars, 200 μm.

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Migration, Mouse Assay

    Cohort of cells labeled at E17.0. A-C: Coronal section of P1.0 (A) and P5.0 (B) brains labeled at E17.0. See also Figure S8 for lower magnification pictures of E17.5 through P5. Higher magnification pictures of E17.5 through P5 from the dorsal cortex were shown in (C). At E17.5, most of the labeled cells were located in the VZ (C). At E18.0, most of the labeled cells were located in the VZ and MAZ (C). Small number of labeled cells were also found throughout the cortex sparsely. At E18.5, many labeled cells were in the MAZ (C). Some labeled cells sparsely distributed throughout the cortex. At E19.0, many cells entered the L1-positive IZ dorsally (C). Small number of cells were also found in the MZ and CP (F). At P1.0, many labeled cells were migrating in the IZ / white matter (A, C). Migrating cells formed a slightly dense cellular structure (inset in A) sandwiched by L1-positive axon bundles (arrowheads in A). At P2.0, many neurons were migrating in the CP/cortical gray matter with a locomotion morphology (C). At P3.0, many labeled cells reached the dorsal PCZ (C). At P5.0, most of the labeled cells were located in the most superficial part of the cortical gray matter (B, C). Note that many cells were located in the dorsal (and dorsolateral) cortex (the yellow dotted line), and few cells were located in the dorsomedial and lateral cortices. D-F : Analyses of GABAergic interneurons. Cells labeled with FT (CytoTell Blue) at E17.0 sparsely distributed throughout the cortex at E18.0 (D, E), and they were mostly positive for GFP in GAD67-GFP mice (E). Labeled cells with similar morphologies were found in the MZ / Layer I and CP at E19.0 before the main population of the labeled cells reach the CP (F). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone; PSB, pallial-subpallial boundary; LI, cortical layer I; GM, gray matter; WM, white matter. Scale bars, 200 μm (A, B, D), 50 μm (C, E), 10 μm (F). * indicated another brain on the same slide glass.

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Cohort of cells labeled at E17.0. A-C: Coronal section of P1.0 (A) and P5.0 (B) brains labeled at E17.0. See also Figure S8 for lower magnification pictures of E17.5 through P5. Higher magnification pictures of E17.5 through P5 from the dorsal cortex were shown in (C). At E17.5, most of the labeled cells were located in the VZ (C). At E18.0, most of the labeled cells were located in the VZ and MAZ (C). Small number of labeled cells were also found throughout the cortex sparsely. At E18.5, many labeled cells were in the MAZ (C). Some labeled cells sparsely distributed throughout the cortex. At E19.0, many cells entered the L1-positive IZ dorsally (C). Small number of cells were also found in the MZ and CP (F). At P1.0, many labeled cells were migrating in the IZ / white matter (A, C). Migrating cells formed a slightly dense cellular structure (inset in A) sandwiched by L1-positive axon bundles (arrowheads in A). At P2.0, many neurons were migrating in the CP/cortical gray matter with a locomotion morphology (C). At P3.0, many labeled cells reached the dorsal PCZ (C). At P5.0, most of the labeled cells were located in the most superficial part of the cortical gray matter (B, C). Note that many cells were located in the dorsal (and dorsolateral) cortex (the yellow dotted line), and few cells were located in the dorsomedial and lateral cortices. D-F : Analyses of GABAergic interneurons. Cells labeled with FT (CytoTell Blue) at E17.0 sparsely distributed throughout the cortex at E18.0 (D, E), and they were mostly positive for GFP in GAD67-GFP mice (E). Labeled cells with similar morphologies were found in the MZ / Layer I and CP at E19.0 before the main population of the labeled cells reach the CP (F). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone; PSB, pallial-subpallial boundary; LI, cortical layer I; GM, gray matter; WM, white matter. Scale bars, 200 μm (A, B, D), 50 μm (C, E), 10 μm (F). * indicated another brain on the same slide glass.

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling, Mouse Assay

    Cohort of cells born at E14.5. A-J: Coronal sections of E15.0 (A), 15.5 (B), 16.0 (C), 16.5 (D), 17.5 (E), 18.5 (F) and P0.5 (G) brains labeled at E14.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex were shown in (H) and (I), respectively. Higher magnification of the apical part of the dorsolateral cortical wall of E15.0 (0.5 days after injection) brains was shown in (J). At E15.0, most of the labeled cells were located in the VZ in both dorsomedial and dorsolateral cortex (A, H, I). Some labeled cells were located outside of the VZ in the dorsolateral cortex (A, I, J) but such cells were not frequently found in the dorsomedial cortex (A, H). The labeled cells that were located basally often had a long ascending process (red arrowheads, J, left) as well as some retraction bulb (blue arrowheads) and were immunoreactive for Pax6, Sox2 and Ki-67 (yellow arrowheads, J, right). Note that the ascending processes were so long that it was difficult to observe full length in the IZ crowded with radial fibers, which are also labeled with FT. At E15.5, majority of the labeled neurons were located in the MAZ in the multipolar morphology in both the dorsomedial and dorsolateral cortex (B, H, I). At E16.0, most of the labeled cells were in the IZ (C, H, I). At E16.5 in the dorsomedial cortex, many cells reached the most superficial part of the CP (D, H). In the dorsolateral CP, on the other hand, most of the labeled cells were migrating in the IZ just beneath the SP (D, I; see also Figures 2A, C). At E17.5 in the dorsomedial cortex, vast majority of the labeled cells were located in the primitive cortical zone (PCZ), which is the most superficial part of the CP (E, H). In the dorsolateral cortex, most of the labeled cells were still migrating in the CP (E, I). At E18.5 in the dorsomedial cortex, labeled cells were distributed not only in the PCZ, but also in the slightly deeper part of the CP as NeuN-positive mature neurons (F, H). In the dorsolateral cortex, majority of the labeled cells were located in the PCZ (F, I). At P0.5 in the dorsolateral cortex, many labeled cells were distributed in the slightly deeper part of the CP as NeuN-positive mature neurons (G). Small double headed arrows show PCZ. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone. Scale bars, 200 μm (A-G), 50 μm (H, I), 10 μm (J).

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Cohort of cells born at E14.5. A-J: Coronal sections of E15.0 (A), 15.5 (B), 16.0 (C), 16.5 (D), 17.5 (E), 18.5 (F) and P0.5 (G) brains labeled at E14.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex were shown in (H) and (I), respectively. Higher magnification of the apical part of the dorsolateral cortical wall of E15.0 (0.5 days after injection) brains was shown in (J). At E15.0, most of the labeled cells were located in the VZ in both dorsomedial and dorsolateral cortex (A, H, I). Some labeled cells were located outside of the VZ in the dorsolateral cortex (A, I, J) but such cells were not frequently found in the dorsomedial cortex (A, H). The labeled cells that were located basally often had a long ascending process (red arrowheads, J, left) as well as some retraction bulb (blue arrowheads) and were immunoreactive for Pax6, Sox2 and Ki-67 (yellow arrowheads, J, right). Note that the ascending processes were so long that it was difficult to observe full length in the IZ crowded with radial fibers, which are also labeled with FT. At E15.5, majority of the labeled neurons were located in the MAZ in the multipolar morphology in both the dorsomedial and dorsolateral cortex (B, H, I). At E16.0, most of the labeled cells were in the IZ (C, H, I). At E16.5 in the dorsomedial cortex, many cells reached the most superficial part of the CP (D, H). In the dorsolateral CP, on the other hand, most of the labeled cells were migrating in the IZ just beneath the SP (D, I; see also Figures 2A, C). At E17.5 in the dorsomedial cortex, vast majority of the labeled cells were located in the primitive cortical zone (PCZ), which is the most superficial part of the CP (E, H). In the dorsolateral cortex, most of the labeled cells were still migrating in the CP (E, I). At E18.5 in the dorsomedial cortex, labeled cells were distributed not only in the PCZ, but also in the slightly deeper part of the CP as NeuN-positive mature neurons (F, H). In the dorsolateral cortex, majority of the labeled cells were located in the PCZ (F, I). At P0.5 in the dorsolateral cortex, many labeled cells were distributed in the slightly deeper part of the CP as NeuN-positive mature neurons (G). Small double headed arrows show PCZ. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone. Scale bars, 200 μm (A-G), 50 μm (H, I), 10 μm (J).

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling, Injection

    Cohort of cells labeled at E15.5. A-B: Coronal sections of E16.0, 16.5, 17.0, 17.5, 18.5, P0.5 and P1.5 brains labeled at E15.5. Data from the dorsomedial cortex and dorsolateral cortex were shown in (A) and (B), respectively. See Figure S7 for low magnification pictures. Figure S7 shows the same sections shown with immunohistochemistry helpful to define histology. At E16.0, most of the labeled cells were in the VZ (A, B, Figures S7A, S7H, S7I). Some labeled cells, often positive for Pax6, were outside of the VZ (B, Figure S7A, S7I) (arrowheads). One day (E16.5; A, B, Figures S7B, S7H, S7I) and 1.5-2 days (E17.0-17.5; A, B, Figures S6C, S6D, S6H, S6I) after injection, most of the labeled cells were in the MAZ and IZ, respectively. At E17.5, most of the labeled cells were migrating in the superficial and deep part of the IZ in the dorsomedial cortex (A, Figures S7D, S7H). In the dorsolateral cortex, migrating cells were mainly in the rather deep part of the IZ (B, Figure S7D, S7I). At E18.5 in the dorsomedial cortex, most of the labeled cells the PCZ (A, Figure S7E, S7H). In the dorsolateral cortex, on the other hand, only the small population of the labeled cells reached the PCZ and others were still migrating in the CP and SP in a locomotion morphology (B, Figures S7E, I). At P0.5, vast majority of the labeled cells settled in the PCZ in the dorsomedial cortex (A, Figures S7F, S7H). In the dorsolateral cortex, too, many labeled cells reached the PCZ (B, Figures S7F, I). At P1.5, labeled cells labeled at E15.5 settled in the gray matter in the dorsolateral cortex (B, Figures S7G, H, I). Some of these labeled cells changed their position slightly apically to leave from the PCZ in the dorsolateral cortex (B, Figures S7G, H, I). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone; PSB, pallial-subpallial boundary; LI, cortical layer I (L1 in the IZ is an axonal marker); GM, gray matter; WM, white matter. Scale bars, 50 μm.

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Cohort of cells labeled at E15.5. A-B: Coronal sections of E16.0, 16.5, 17.0, 17.5, 18.5, P0.5 and P1.5 brains labeled at E15.5. Data from the dorsomedial cortex and dorsolateral cortex were shown in (A) and (B), respectively. See Figure S7 for low magnification pictures. Figure S7 shows the same sections shown with immunohistochemistry helpful to define histology. At E16.0, most of the labeled cells were in the VZ (A, B, Figures S7A, S7H, S7I). Some labeled cells, often positive for Pax6, were outside of the VZ (B, Figure S7A, S7I) (arrowheads). One day (E16.5; A, B, Figures S7B, S7H, S7I) and 1.5-2 days (E17.0-17.5; A, B, Figures S6C, S6D, S6H, S6I) after injection, most of the labeled cells were in the MAZ and IZ, respectively. At E17.5, most of the labeled cells were migrating in the superficial and deep part of the IZ in the dorsomedial cortex (A, Figures S7D, S7H). In the dorsolateral cortex, migrating cells were mainly in the rather deep part of the IZ (B, Figure S7D, S7I). At E18.5 in the dorsomedial cortex, most of the labeled cells the PCZ (A, Figure S7E, S7H). In the dorsolateral cortex, on the other hand, only the small population of the labeled cells reached the PCZ and others were still migrating in the CP and SP in a locomotion morphology (B, Figures S7E, I). At P0.5, vast majority of the labeled cells settled in the PCZ in the dorsomedial cortex (A, Figures S7F, S7H). In the dorsolateral cortex, too, many labeled cells reached the PCZ (B, Figures S7F, I). At P1.5, labeled cells labeled at E15.5 settled in the gray matter in the dorsolateral cortex (B, Figures S7G, H, I). Some of these labeled cells changed their position slightly apically to leave from the PCZ in the dorsolateral cortex (B, Figures S7G, H, I). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone; PSB, pallial-subpallial boundary; LI, cortical layer I (L1 in the IZ is an axonal marker); GM, gray matter; WM, white matter. Scale bars, 50 μm.

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling, Immunohistochemistry, Injection, Marker

    Cohort of cells born at E10.5. A-E: Coronal sections of 12.5 (A), 13.5 (B) and 16.5 (C) brains labeled at E10.5. See also Figure S2 for coronal section from E11.5 to 16.5 shown with FT and DAPI. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex from E11.5 to 16.5 were shown in (D) and (E), respectively. As early as E11.5, some cells were found in the preplate (PP), which was very thin in the dorsomedial cortex, as well as in the VZ (D, E, Figure S2A). At E12.5, many cells were in the PP, sometimes in a tangential morphology (A, D, E). At E13.5, the CSPG and nuclear staining showed PP splitting proceeded in a lateral-to-medial direction and the CP (asterisks) was observed in the dorsolateral cortex but not in the dorsomedial cortex (B). In the dorsomedial cortex, labeled cells were in the PP, often in somewhat round morphology (D). In the dorsolateral cortex, on the other hand, many labeled cells were located in the CP (shown with blue arrows) and MZ (E). Note that few cells were found below the CP identified by nuclear and CSPG staining (B, E, Figure S2C). At E14.5, thin CP was identified in the dorsomedial cortex as well (D, E, Figure S2D). Some labeled cells were in the deep part of the CP in the dorsomedial cortex, but many labeled cells were still in the MZ (D). In the dorsolateral cortex, many labeled cells were found near the boundary between the CP and SP (D, Figure S2D). At E15.5, labeled cells were found at the boundary between SP and CP as well as MZ in the dorsomedial cortex (D, Figure S2E, S2E’), which is similar to the dorsolateral cortex of the E14.5 (E, Figure S2D). In the E15.5 dorsolateral cortex, many labeled cells were in the CSPG-positive SP (D, E, Figure S2E, S2E’). At E16.5, in both the dorsomedial and dorsolateral cortex, labeled cells were mainly found in the SP (C, D, E). Some cells were also found in the MZ (D, E, Figure S2G). Note that the CSPG staining in the SP showed some double-track immunoreactivity strong just above and below the distinct cell layer in the SP in dorsal and dorsolateral cortex at E15.5-E16.5 (E). Emergence of the labeled cells in the SP seems to coincide with this emergence of distinct layer. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Cohort of cells born at E10.5. A-E: Coronal sections of 12.5 (A), 13.5 (B) and 16.5 (C) brains labeled at E10.5. See also Figure S2 for coronal section from E11.5 to 16.5 shown with FT and DAPI. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex from E11.5 to 16.5 were shown in (D) and (E), respectively. As early as E11.5, some cells were found in the preplate (PP), which was very thin in the dorsomedial cortex, as well as in the VZ (D, E, Figure S2A). At E12.5, many cells were in the PP, sometimes in a tangential morphology (A, D, E). At E13.5, the CSPG and nuclear staining showed PP splitting proceeded in a lateral-to-medial direction and the CP (asterisks) was observed in the dorsolateral cortex but not in the dorsomedial cortex (B). In the dorsomedial cortex, labeled cells were in the PP, often in somewhat round morphology (D). In the dorsolateral cortex, on the other hand, many labeled cells were located in the CP (shown with blue arrows) and MZ (E). Note that few cells were found below the CP identified by nuclear and CSPG staining (B, E, Figure S2C). At E14.5, thin CP was identified in the dorsomedial cortex as well (D, E, Figure S2D). Some labeled cells were in the deep part of the CP in the dorsomedial cortex, but many labeled cells were still in the MZ (D). In the dorsolateral cortex, many labeled cells were found near the boundary between the CP and SP (D, Figure S2D). At E15.5, labeled cells were found at the boundary between SP and CP as well as MZ in the dorsomedial cortex (D, Figure S2E, S2E’), which is similar to the dorsolateral cortex of the E14.5 (E, Figure S2D). In the E15.5 dorsolateral cortex, many labeled cells were in the CSPG-positive SP (D, E, Figure S2E, S2E’). At E16.5, in both the dorsomedial and dorsolateral cortex, labeled cells were mainly found in the SP (C, D, E). Some cells were also found in the MZ (D, E, Figure S2G). Note that the CSPG staining in the SP showed some double-track immunoreactivity strong just above and below the distinct cell layer in the SP in dorsal and dorsolateral cortex at E15.5-E16.5 (E). Emergence of the labeled cells in the SP seems to coincide with this emergence of distinct layer. DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-C) and 50 μm (D, E).

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling, Staining

    Characterization of cell population labeled with the FlashTag (FT) technology. A-G: 1 mM of 5- or 6-(N-Succinimidyloxycarbonyl) fluorescein 3’,6’-diacetate (CFSE) was injected into the lateral ventricles (LV) at embryonic day (E)14 ICR mice and fixed 0.5 (A), 3.5 (B), 6.5 (C), 9.5 (D) hours later. Intraperitoneal bolus injection of 5-Ethynyl-2’-deoxyuridine (EdU) was performed maternally 0.5 hours before fixation. Photomicrographs from the dorsolateral cortex were shown. In (A), FT-labeled cells positioned most apically and were often positive for phospho-histone H3 (pH3) ( Figure 1F -0.5 hour, dorsolateral: 36.1± 4.0% [mean ± standard error of means], 339 cells from 5 brains; dorsomedial: 35.2 ± 4.7%, 249 cells from 5 brains) but negative for EdU administered at the same time ( Figure 1G , -0.5 hour, dorsolateral: 0 ± 0%, 339 cells from 5 brains; dorsomedial: 0 ± 0%, 249 cells from 5 brains). The nuclei of EdU positive cells positioned basally in the VZ. 3.5 hours after FT injection, FT-labeled cells left the ventricular surface but still near it, and were no longer positive for pH3 (B) ( Figure 1F , -3.5 hours, dorsolateral: 2.8 ± 0.7%, 530 cells from 5 brains; dorsomedial: 1.1 ± 0.5%, 415 cells from 5 brains). 6.5 hours after labeling, almost no cells were adjacent to the lateral ventricle (C). 9.5 hours after labeling, most of the labeled cells were in about basal two thirds in the VZ and double labeled for EdU, suggesting that some of them reentered the S-phase (D) ( Figure 1G , -9.5 hours, dorsolateral: 15.9± 2.5%, 711 cells from 5 brains; dorsomedial: 33.4 ± 6.6 %, 546 cells from 5 brains). Schematic presentation of these experiments was shown in E. In (F), percentages of pH3+ cells out of the FT-labeled cells were shown. Magenta, pH3+ FT+/FT+ in the dorsolateral cortex. Green, pH3+ FT+/FT+ in the dorsomedial cortex. In (G), percentages of EdU+ cells out of FT-labeled cells were shown. Orange, EdU+ FT+/FT+ in the dorsolateral cortex. Blue, EdU+ FT+/FT+ in the dorsomedial cortex. H-L: EdU was administered 3 (I), 6 (J) and 9 (K) hours before FT labeling. 0.5 hours after FT labeling, brains were harvested. Schematic presentation of these experiments was shown in (H). Nuclei of the EdU labeled cells positioned more apically in brains in which EdU was administered 3.5 hours before fixation (I) compared with A, and some of the EdU labeled cells positioned at the ventricular surface to enter M phase (interkinetic nuclear migration). In brains which EdU was administered 6.5 (J) and 9.5 (K) hours before fixation, EdU-labeled cells positioned even more apically. In these mice treated with EdU 3-9 hours prior to FT, FT-labeled cells were often co-labelled with EdU (I-K) ( Figure1L , dorsolateral, -9.5 hours: 76.6 ± 2.4%, 328 cells from 5 brains; -6.5 hours: 96.1 ± 0.5 %, 304 cells from 5 brains; -3.5 hours: 81.2 ± 1.9 %, 263 cells from 5 brains. Dorsomedial, -9.5 hours: 65.1 ± 1.5%, 369 cells from 5 brains; -6.5 hours: 96.7 ± 1.2 %, 217 cells from 5 brains; -3.5 hours: 81.5 ± 1.9 %, 287 cells from 5 brains). Note that EdU and FT never co-labeled when administered simultaneously (A). In the graph in (L), percentage of EdU+ cells out of FT-labeled cells were shown. Data for -0.5 hours in (L) corresponds to those for -0.5 hours in (G). Orange, EdU+ FT+/FT+ in the dorsolateral cortex. Blue, EdU+ FT+/FT+ in the dorsomedial cortex. M-O: CytoTell Blue was injected into the LV of the E12.5 (M-N) and 15.5 (O) GAD67-GFP brains. In E15.5 dorsolateral cortex labeled at E12.5, most of the labeled cells (red) were in the deep part of the cortical plate (CP) (M, N). Vast majority of the labeled cells were negative for GFP (E12.5-15.5 dorsolateral cortex, 93.3 ± 2.5%, 1653 cells from 3 brains) (N, N1-3). In postnatal day (P)1 dorsolateral cortex labeled at E15.5 (O), most of the labeled cells were found in the superficial gray matter. Again, vast majority of the labeled cells were negative for GFP (E15.5-P1, 95.5 ± 0.5 %, 1455 cells from 5 brains) (O, O1-3). Arrowheads in (N) and (O) show rare examples of cells positive for both FT and GFP. EdU, 5-Ethynyl-2’-deoxyuridine; DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; pH3, phospho-histone H3. GAD-GFP, Glutamate decarboxylase 67-green fluorescent protein, VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone; LI, cortical layer I; GM, gray matter; WM, white matter. Scale bars, 20 μm (A-D, I-K), 50 μm (N, O), 200 μm (M).

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Characterization of cell population labeled with the FlashTag (FT) technology. A-G: 1 mM of 5- or 6-(N-Succinimidyloxycarbonyl) fluorescein 3’,6’-diacetate (CFSE) was injected into the lateral ventricles (LV) at embryonic day (E)14 ICR mice and fixed 0.5 (A), 3.5 (B), 6.5 (C), 9.5 (D) hours later. Intraperitoneal bolus injection of 5-Ethynyl-2’-deoxyuridine (EdU) was performed maternally 0.5 hours before fixation. Photomicrographs from the dorsolateral cortex were shown. In (A), FT-labeled cells positioned most apically and were often positive for phospho-histone H3 (pH3) ( Figure 1F -0.5 hour, dorsolateral: 36.1± 4.0% [mean ± standard error of means], 339 cells from 5 brains; dorsomedial: 35.2 ± 4.7%, 249 cells from 5 brains) but negative for EdU administered at the same time ( Figure 1G , -0.5 hour, dorsolateral: 0 ± 0%, 339 cells from 5 brains; dorsomedial: 0 ± 0%, 249 cells from 5 brains). The nuclei of EdU positive cells positioned basally in the VZ. 3.5 hours after FT injection, FT-labeled cells left the ventricular surface but still near it, and were no longer positive for pH3 (B) ( Figure 1F , -3.5 hours, dorsolateral: 2.8 ± 0.7%, 530 cells from 5 brains; dorsomedial: 1.1 ± 0.5%, 415 cells from 5 brains). 6.5 hours after labeling, almost no cells were adjacent to the lateral ventricle (C). 9.5 hours after labeling, most of the labeled cells were in about basal two thirds in the VZ and double labeled for EdU, suggesting that some of them reentered the S-phase (D) ( Figure 1G , -9.5 hours, dorsolateral: 15.9± 2.5%, 711 cells from 5 brains; dorsomedial: 33.4 ± 6.6 %, 546 cells from 5 brains). Schematic presentation of these experiments was shown in E. In (F), percentages of pH3+ cells out of the FT-labeled cells were shown. Magenta, pH3+ FT+/FT+ in the dorsolateral cortex. Green, pH3+ FT+/FT+ in the dorsomedial cortex. In (G), percentages of EdU+ cells out of FT-labeled cells were shown. Orange, EdU+ FT+/FT+ in the dorsolateral cortex. Blue, EdU+ FT+/FT+ in the dorsomedial cortex. H-L: EdU was administered 3 (I), 6 (J) and 9 (K) hours before FT labeling. 0.5 hours after FT labeling, brains were harvested. Schematic presentation of these experiments was shown in (H). Nuclei of the EdU labeled cells positioned more apically in brains in which EdU was administered 3.5 hours before fixation (I) compared with A, and some of the EdU labeled cells positioned at the ventricular surface to enter M phase (interkinetic nuclear migration). In brains which EdU was administered 6.5 (J) and 9.5 (K) hours before fixation, EdU-labeled cells positioned even more apically. In these mice treated with EdU 3-9 hours prior to FT, FT-labeled cells were often co-labelled with EdU (I-K) ( Figure1L , dorsolateral, -9.5 hours: 76.6 ± 2.4%, 328 cells from 5 brains; -6.5 hours: 96.1 ± 0.5 %, 304 cells from 5 brains; -3.5 hours: 81.2 ± 1.9 %, 263 cells from 5 brains. Dorsomedial, -9.5 hours: 65.1 ± 1.5%, 369 cells from 5 brains; -6.5 hours: 96.7 ± 1.2 %, 217 cells from 5 brains; -3.5 hours: 81.5 ± 1.9 %, 287 cells from 5 brains). Note that EdU and FT never co-labeled when administered simultaneously (A). In the graph in (L), percentage of EdU+ cells out of FT-labeled cells were shown. Data for -0.5 hours in (L) corresponds to those for -0.5 hours in (G). Orange, EdU+ FT+/FT+ in the dorsolateral cortex. Blue, EdU+ FT+/FT+ in the dorsomedial cortex. M-O: CytoTell Blue was injected into the LV of the E12.5 (M-N) and 15.5 (O) GAD67-GFP brains. In E15.5 dorsolateral cortex labeled at E12.5, most of the labeled cells (red) were in the deep part of the cortical plate (CP) (M, N). Vast majority of the labeled cells were negative for GFP (E12.5-15.5 dorsolateral cortex, 93.3 ± 2.5%, 1653 cells from 3 brains) (N, N1-3). In postnatal day (P)1 dorsolateral cortex labeled at E15.5 (O), most of the labeled cells were found in the superficial gray matter. Again, vast majority of the labeled cells were negative for GFP (E15.5-P1, 95.5 ± 0.5 %, 1455 cells from 5 brains) (O, O1-3). Arrowheads in (N) and (O) show rare examples of cells positive for both FT and GFP. EdU, 5-Ethynyl-2’-deoxyuridine; DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; pH3, phospho-histone H3. GAD-GFP, Glutamate decarboxylase 67-green fluorescent protein, VZ, ventricular zone; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; PCZ, primitive cortical zone; MZ, marginal zone; LI, cortical layer I; GM, gray matter; WM, white matter. Scale bars, 20 μm (A-D, I-K), 50 μm (N, O), 200 μm (M).

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling, Injection, Mouse Assay, Migration

    Cohort of cells born at E13.5. A-G: Coronal sections of E14.0 (A), 14.5 (B), 15.0 (C), 15.5 (D) and 16.5 (E) brains labeled at E13.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex from E14.0-17.5 were shown in (F) and (G), respectively. Lower magnification pictures of E17.5 and E18.5 are shown in Figure S5. Coronal sections of FT-labeled brains of E14.0 through E18.5 stained with nuclear staining were also shown in Figure S5. At E14.0, most of the labeled cells were located in the VZ and zones just above the VZ in both dorsomedial and dorsolateral cortex (A, F, G). At E14.5, many labeled neurons were migrating in the IZ below the SP as revealed by immunohistochemistry for CSPG (B, F, G). At E15.0, majority of the labeled cells reached just beneath the pial surface in the dorsomedial cortex (C, F) but most of the labeled cells in the dorsolateral cortex were in the IZ below the SP (C, G). At E15.5, some of the labeled cells entered the CP while many neurons were still migrating in the IZ and SP in the dorsolateral cortex (D, G). At E16.5, most of the labeled cells in the dorsomedial cortex were located in the CP (E, F). Most of the labeled cells in the dorsolateral cortex reached the superficial part of the CP. Note that FT-labeled axon bundles were in the IZ. At E17.5, many strongly labeled neurons located in the deeper part of the CP, suggesting that later-born neurons passed through the neuronal layers that were born at E13.5 (F, G, Figure S5F, S5G). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-E), 50 μm (F, G).

    Journal: bioRxiv

    Article Title: Comprehensive characterization of migration profiles of murine cerebral cortical neurons during development using FlashTag labeling

    doi: 10.1101/2020.10.05.317925

    Figure Lengend Snippet: Cohort of cells born at E13.5. A-G: Coronal sections of E14.0 (A), 14.5 (B), 15.0 (C), 15.5 (D) and 16.5 (E) brains labeled at E13.5. Higher magnification pictures from the dorsomedial cortex and dorsolateral cortex from E14.0-17.5 were shown in (F) and (G), respectively. Lower magnification pictures of E17.5 and E18.5 are shown in Figure S5. Coronal sections of FT-labeled brains of E14.0 through E18.5 stained with nuclear staining were also shown in Figure S5. At E14.0, most of the labeled cells were located in the VZ and zones just above the VZ in both dorsomedial and dorsolateral cortex (A, F, G). At E14.5, many labeled neurons were migrating in the IZ below the SP as revealed by immunohistochemistry for CSPG (B, F, G). At E15.0, majority of the labeled cells reached just beneath the pial surface in the dorsomedial cortex (C, F) but most of the labeled cells in the dorsolateral cortex were in the IZ below the SP (C, G). At E15.5, some of the labeled cells entered the CP while many neurons were still migrating in the IZ and SP in the dorsolateral cortex (D, G). At E16.5, most of the labeled cells in the dorsomedial cortex were located in the CP (E, F). Most of the labeled cells in the dorsolateral cortex reached the superficial part of the CP. Note that FT-labeled axon bundles were in the IZ. At E17.5, many strongly labeled neurons located in the deeper part of the CP, suggesting that later-born neurons passed through the neuronal layers that were born at E13.5 (F, G, Figure S5F, S5G). DAPI, 4’,6-diamidino-2-phenylindole; FT, FlashTag; VZ, ventricular zone; PP, preplate; MAZ, multipolar cell accumulation zone; IZ, intermediate zone; SP, subplate; CP, cortical plate; MZ, marginal zone. Scale bars, 200 μm (A-E), 50 μm (F, G).

    Article Snippet: When nuclear staining was performed without immunohistochemistry, sections were immersed with PBS for more than 30 minutes at RT and incubated with 2.5 ng/μl of 4’,6-diamidino-2-phenylindole (DAPI; D3571; Thermo Fisher Scientific, Waltham, MA) or 0.5 μM of TO-PRO3 Iodide (T3605, Thermo Fisher Scientific) at RT for 1 hour.

    Techniques: Labeling, Staining, Immunohistochemistry

    Akt2 promotes NKT17 lineage differentiation. Cells were from WT and Akt2 KO mice. (A) Intracellular staining of PLZF, RORγt, T-bet, and GATA3 in iNKT cells from the thymus. Percentages and absolute numbers of NKT1, NKT2, NKT17 cells in the thymus ( n = 6 mice per group) (B) and spleen ( n = 4 mice per group) (C) from WT, KO, WT chimera mice ( n = 3) and Akt2 KO chimera mice ( n = 3). (D–F) Cytokine production by iNKT cells (gated on CD1d-PBS57 + TCRβ + cells) from the thymus ( n = 5 mice per group) and spleen ( n = 5 mice per group) after stimulation with PMA plus ionomycin for 5 h. (G,H) Cytokine production by iNKT cells from WT and Akt2 KO thymocytes stimulated with α-GalCer for 72 h and with PMA plus ionomycin in the last 5 h ( n = 4 mice per group). (I) Critical role of Akt2 for PLZF localization to the nuclear bodies. MACS—enriched and FACS—sorted NKT17 cells from WT and Akt2 KO thymocytes( n = 3) were fixed and stained with a mouse anti-PLZF and Actin as primary antibody, detected with an Goat anti mouse secondary antibody. The nuclei were stained with DAPI. (J) Mean fluorescent intensity of cytoplasm and nucleus. * p

    Journal: Frontiers in Immunology

    Article Title: Akt2 Regulates the Differentiation and Function of NKT17 Cells via FoxO-1-ICOS Axis

    doi: 10.3389/fimmu.2018.01940

    Figure Lengend Snippet: Akt2 promotes NKT17 lineage differentiation. Cells were from WT and Akt2 KO mice. (A) Intracellular staining of PLZF, RORγt, T-bet, and GATA3 in iNKT cells from the thymus. Percentages and absolute numbers of NKT1, NKT2, NKT17 cells in the thymus ( n = 6 mice per group) (B) and spleen ( n = 4 mice per group) (C) from WT, KO, WT chimera mice ( n = 3) and Akt2 KO chimera mice ( n = 3). (D–F) Cytokine production by iNKT cells (gated on CD1d-PBS57 + TCRβ + cells) from the thymus ( n = 5 mice per group) and spleen ( n = 5 mice per group) after stimulation with PMA plus ionomycin for 5 h. (G,H) Cytokine production by iNKT cells from WT and Akt2 KO thymocytes stimulated with α-GalCer for 72 h and with PMA plus ionomycin in the last 5 h ( n = 4 mice per group). (I) Critical role of Akt2 for PLZF localization to the nuclear bodies. MACS—enriched and FACS—sorted NKT17 cells from WT and Akt2 KO thymocytes( n = 3) were fixed and stained with a mouse anti-PLZF and Actin as primary antibody, detected with an Goat anti mouse secondary antibody. The nuclei were stained with DAPI. (J) Mean fluorescent intensity of cytoplasm and nucleus. * p

    Article Snippet: The unstimulated iNKT cells were incubated with a mouse anti-PLZF antibody (4 μg/ml, Santa Cruz Biotechnology) and AF488-Actin (Invitrogen) for 1 h, further stained with a Goat anti-mouse AF546 secondary antibody (1:400) for 30 min, and finally covered with 1.5 μg/ml DAPI (Beyotime). α-GalCer stimulated iNTK cells were incubated with rabbit-anti-FoxO-1 (Cell signaling technology) and AF488-Actin for 1 h, and further stained with a AF546-Goat anti-rabbit secondary antibody (1:400) for 30 min, and finally covered with 1.5 μg/ml DAPI (Beyotime).

    Techniques: Mouse Assay, Staining, Magnetic Cell Separation, FACS

    Akt2 couples with FoxO-1 to regulate the expression of ICOS in NKT cells. (A) MACS—enriched and FACS—sorted iNKT cells from WT and Akt2 KO thymocytes were fixed and stained with a rabbit-anti-FoxO1 and AF488-Actin as primary antibody, detected with a goat anti rabbit secondary antibody. The nuclei were stained with DAPI. (B) MACS—enriched and FACS—sorted iNKT cells from WT and KO thymocytes were incubated with125 ng/mL α-GalCer for 3 d and then stained with FoxO1 and AF488-Actin as primary antibody, detected with a Goat anti mouse secondary antibody. (C) MFI of FoxO1 of iNKT cells in the thymus from the WT and Akt2 KO mice ( n = 3) quantified with NIS-Elements AR 3.2 software. (D) Phospho flow analysis of the MFI of pFoxO-1(S256) in the WT and KO thymocytes. (E) MACS—enriched and FACS—sorted iNKT cells from WT and ICOS KO thymocytes were fixed and stained with a rabbit-anti-FoxO1 and AF488-Actin as primary antibody, detected with a goat anti rabbit secondary antibody. The nuclei were stained with DAPI. (F) MACS—–enriched and FACS—sorted iNKT cells from WT and ICOS KO thymocytes ( n = 3 mice per group) were incubated with125 ng/mL α-GalCer for 3 d and then stained with FoxO1 and AF488-Actin as primary antibody, detected with a Goat anti mouse secondary antibody. (G) MFI of FoxO1 of iNKT cells in the thymus from the WT ( n = 3) and ICOS KO mice ( n = 3) quantified with NIS-Elements AR 3.2 software. (H) Phospho flow analysis of the MFI of pFoxO-1(S256) in the WT and ICOS KO thymocytes from three independent experiments. * p

    Journal: Frontiers in Immunology

    Article Title: Akt2 Regulates the Differentiation and Function of NKT17 Cells via FoxO-1-ICOS Axis

    doi: 10.3389/fimmu.2018.01940

    Figure Lengend Snippet: Akt2 couples with FoxO-1 to regulate the expression of ICOS in NKT cells. (A) MACS—enriched and FACS—sorted iNKT cells from WT and Akt2 KO thymocytes were fixed and stained with a rabbit-anti-FoxO1 and AF488-Actin as primary antibody, detected with a goat anti rabbit secondary antibody. The nuclei were stained with DAPI. (B) MACS—enriched and FACS—sorted iNKT cells from WT and KO thymocytes were incubated with125 ng/mL α-GalCer for 3 d and then stained with FoxO1 and AF488-Actin as primary antibody, detected with a Goat anti mouse secondary antibody. (C) MFI of FoxO1 of iNKT cells in the thymus from the WT and Akt2 KO mice ( n = 3) quantified with NIS-Elements AR 3.2 software. (D) Phospho flow analysis of the MFI of pFoxO-1(S256) in the WT and KO thymocytes. (E) MACS—enriched and FACS—sorted iNKT cells from WT and ICOS KO thymocytes were fixed and stained with a rabbit-anti-FoxO1 and AF488-Actin as primary antibody, detected with a goat anti rabbit secondary antibody. The nuclei were stained with DAPI. (F) MACS—–enriched and FACS—sorted iNKT cells from WT and ICOS KO thymocytes ( n = 3 mice per group) were incubated with125 ng/mL α-GalCer for 3 d and then stained with FoxO1 and AF488-Actin as primary antibody, detected with a Goat anti mouse secondary antibody. (G) MFI of FoxO1 of iNKT cells in the thymus from the WT ( n = 3) and ICOS KO mice ( n = 3) quantified with NIS-Elements AR 3.2 software. (H) Phospho flow analysis of the MFI of pFoxO-1(S256) in the WT and ICOS KO thymocytes from three independent experiments. * p

    Article Snippet: The unstimulated iNKT cells were incubated with a mouse anti-PLZF antibody (4 μg/ml, Santa Cruz Biotechnology) and AF488-Actin (Invitrogen) for 1 h, further stained with a Goat anti-mouse AF546 secondary antibody (1:400) for 30 min, and finally covered with 1.5 μg/ml DAPI (Beyotime). α-GalCer stimulated iNTK cells were incubated with rabbit-anti-FoxO-1 (Cell signaling technology) and AF488-Actin for 1 h, and further stained with a AF546-Goat anti-rabbit secondary antibody (1:400) for 30 min, and finally covered with 1.5 μg/ml DAPI (Beyotime).

    Techniques: Expressing, Magnetic Cell Separation, FACS, Staining, Incubation, Mouse Assay, Software, Flow Cytometry

    Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for the time courses of 20 hours (A–D), 30 hours (E–H), and 10 days (I–L). The expressions of GFAP, βTubulin, Nestin, and c-Fos were detected in differentiated (B, F, J, D, H, L) and undifferentiated (A, E, I, C, G, K) TH + rBM-MSCs. Nuclei were shown by DAPI in blue staining. Scale bar: 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, 4´,6-diamidino-2-phenylindole.

    Journal: Neurospine

    Article Title: The Effect of Recombinant Tyrosine Hydroxylase Expression on the Neurogenic Differentiation Potency of Mesenchymal Stem Cells

    doi: 10.14245/ns.1836010.005

    Figure Lengend Snippet: Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for the time courses of 20 hours (A–D), 30 hours (E–H), and 10 days (I–L). The expressions of GFAP, βTubulin, Nestin, and c-Fos were detected in differentiated (B, F, J, D, H, L) and undifferentiated (A, E, I, C, G, K) TH + rBM-MSCs. Nuclei were shown by DAPI in blue staining. Scale bar: 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, 4´,6-diamidino-2-phenylindole.

    Article Snippet: Samples were incubated with appropriate secondary antibodies for 25 minutes and covered with mounting medium containing DAPI (4´,6-diamidino-2-phenylindole) (Santa Cruz Biotechnology).

    Techniques: Fluorescence, Staining

    Immunophenotype analysis of TH + rBM-MSCs for neurogenic markers. Representative staining patterns are shown for Nestin (B, D), c-Fos (C, D), GFAP (F, H), β Tubulin (G, H), NF (J, L), TUBB3 (K, L). Nuclei were labeled with DAPI (A, E, I; blue). Only Nestin, GFAP, and TUBB3 staining were found positive. Scale bars represent 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, 4´,6-diamidino-2-phenylindole.

    Journal: Neurospine

    Article Title: The Effect of Recombinant Tyrosine Hydroxylase Expression on the Neurogenic Differentiation Potency of Mesenchymal Stem Cells

    doi: 10.14245/ns.1836010.005

    Figure Lengend Snippet: Immunophenotype analysis of TH + rBM-MSCs for neurogenic markers. Representative staining patterns are shown for Nestin (B, D), c-Fos (C, D), GFAP (F, H), β Tubulin (G, H), NF (J, L), TUBB3 (K, L). Nuclei were labeled with DAPI (A, E, I; blue). Only Nestin, GFAP, and TUBB3 staining were found positive. Scale bars represent 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, 4´,6-diamidino-2-phenylindole.

    Article Snippet: Samples were incubated with appropriate secondary antibodies for 25 minutes and covered with mounting medium containing DAPI (4´,6-diamidino-2-phenylindole) (Santa Cruz Biotechnology).

    Techniques: Staining, Labeling

    Expression of neurotrophic factors, BDNF, and CNTF, in undifferentiated (A–D) and in neurogenic differentiated TH + rBM-MSCs for 30 hours (E–H). Nuclei were shown in blue by DAPI staining. (I) Cell proliferation analysis for neutralized TH + rBM-MSCs, untreated TH + rBM-MSCs, and rBM-MSCs. (J) Expressions of neurogenic markers in neurogenic differentiated/undifferentiated TH + rBM-MSCs were estimated by real-time polymerase chain reaction. TH showed 4 times higher expression level in neurogenic differentiated TH + rBM-MSCs. Scale Bars: 50 µm. BDNF, brain-derived neurotrophic factor; CNTF, ciliary neurotrophic factor; TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, 4´,6-diamidino-2-phenylindole.

    Journal: Neurospine

    Article Title: The Effect of Recombinant Tyrosine Hydroxylase Expression on the Neurogenic Differentiation Potency of Mesenchymal Stem Cells

    doi: 10.14245/ns.1836010.005

    Figure Lengend Snippet: Expression of neurotrophic factors, BDNF, and CNTF, in undifferentiated (A–D) and in neurogenic differentiated TH + rBM-MSCs for 30 hours (E–H). Nuclei were shown in blue by DAPI staining. (I) Cell proliferation analysis for neutralized TH + rBM-MSCs, untreated TH + rBM-MSCs, and rBM-MSCs. (J) Expressions of neurogenic markers in neurogenic differentiated/undifferentiated TH + rBM-MSCs were estimated by real-time polymerase chain reaction. TH showed 4 times higher expression level in neurogenic differentiated TH + rBM-MSCs. Scale Bars: 50 µm. BDNF, brain-derived neurotrophic factor; CNTF, ciliary neurotrophic factor; TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; DAPI, 4´,6-diamidino-2-phenylindole.

    Article Snippet: Samples were incubated with appropriate secondary antibodies for 25 minutes and covered with mounting medium containing DAPI (4´,6-diamidino-2-phenylindole) (Santa Cruz Biotechnology).

    Techniques: Expressing, Staining, Real-time Polymerase Chain Reaction, Derivative Assay

    Differentiation of TH + rBM-MSCs into neuron-like cells. After 30 hours of induction in the neuronal differentiation medium, cells displayed neuron-like morphologies (arrows) (B, C). (A) There was no neuronal differentiation in control culture. Scale bar: A, 200 µm; B, 50 µm; C, 20 µm. Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for distinct time courses of 20 hours (D–G), 30 hours (H–K), and 10 days (L–O). The expressions of FLAG (D, E, H, I, L, M) and TH (F, G, J, K, N, O) were detected in differentiated (E, I, M, F, J, N) and undifferentiated (D, H, L, F, J, N) TH + rBM-MSCs. Nuclei were shown by DAPI in blue staining. Scale bar: D–O, 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; FLAG, protein expression system; DAPI, 4´,6-diamidino-2-phenylindole.

    Journal: Neurospine

    Article Title: The Effect of Recombinant Tyrosine Hydroxylase Expression on the Neurogenic Differentiation Potency of Mesenchymal Stem Cells

    doi: 10.14245/ns.1836010.005

    Figure Lengend Snippet: Differentiation of TH + rBM-MSCs into neuron-like cells. After 30 hours of induction in the neuronal differentiation medium, cells displayed neuron-like morphologies (arrows) (B, C). (A) There was no neuronal differentiation in control culture. Scale bar: A, 200 µm; B, 50 µm; C, 20 µm. Immune fluorescence analysis of neural differentiated TH + rBM-MSCs for distinct time courses of 20 hours (D–G), 30 hours (H–K), and 10 days (L–O). The expressions of FLAG (D, E, H, I, L, M) and TH (F, G, J, K, N, O) were detected in differentiated (E, I, M, F, J, N) and undifferentiated (D, H, L, F, J, N) TH + rBM-MSCs. Nuclei were shown by DAPI in blue staining. Scale bar: D–O, 50 µm. TH, tyrosine hydroxylase; rBM-MSC, rat bone marrow mesenchymal stem cells; FLAG, protein expression system; DAPI, 4´,6-diamidino-2-phenylindole.

    Article Snippet: Samples were incubated with appropriate secondary antibodies for 25 minutes and covered with mounting medium containing DAPI (4´,6-diamidino-2-phenylindole) (Santa Cruz Biotechnology).

    Techniques: Fluorescence, Staining, Expressing

    The unfolded protein response (UPR) is an early event during in vitro hepatic stellate cell (HSC) activation. a Bright-field images of freshly isolated mouse HSCs (mHSCs), plated on plastic culture dishes, were taken after 2, 10 and 24 h and 4, 7 and 10 days in culture (upper right panel). At the same time points, expression of HSC activation markers Acta2 and Lox was determined at the mRNA level by quantitative real-time polymerase chain reaction (qPCR; upper left panel). The lower panel shows mRNA levels of UPR genes Bip , Chop , Xbp1s , Atf4 and Herpud1 at the indicated time points during HSC culture; n = 3. b Immunofluorescent staining for Bip (green) of HSCs fixed at different time points during cultivation. 4′,6-Diamidino-2-phenylindole (DAPI; blue) was used as a nuclear staining. Graph representing the quantification of % mean intensity in at least 25 cells for each time point. c Western blot analysis of HSC activation markers PDGFRβ and α-smooth muscle actin (α-SMA) and of UPR marker BIP. GAPDH was used as a loading control. Cells were collected at the indicated time points, and samples for the 0 h time point were not seeded, but lysed immediately after cell isolation. Images and graphics are representatives of at least two different HSC isolations. * P

    Journal: Cell Death & Disease

    Article Title: Unfolded protein response is an early, non-critical event during hepatic stellate cell activation

    doi: 10.1038/s41419-019-1327-5

    Figure Lengend Snippet: The unfolded protein response (UPR) is an early event during in vitro hepatic stellate cell (HSC) activation. a Bright-field images of freshly isolated mouse HSCs (mHSCs), plated on plastic culture dishes, were taken after 2, 10 and 24 h and 4, 7 and 10 days in culture (upper right panel). At the same time points, expression of HSC activation markers Acta2 and Lox was determined at the mRNA level by quantitative real-time polymerase chain reaction (qPCR; upper left panel). The lower panel shows mRNA levels of UPR genes Bip , Chop , Xbp1s , Atf4 and Herpud1 at the indicated time points during HSC culture; n = 3. b Immunofluorescent staining for Bip (green) of HSCs fixed at different time points during cultivation. 4′,6-Diamidino-2-phenylindole (DAPI; blue) was used as a nuclear staining. Graph representing the quantification of % mean intensity in at least 25 cells for each time point. c Western blot analysis of HSC activation markers PDGFRβ and α-smooth muscle actin (α-SMA) and of UPR marker BIP. GAPDH was used as a loading control. Cells were collected at the indicated time points, and samples for the 0 h time point were not seeded, but lysed immediately after cell isolation. Images and graphics are representatives of at least two different HSC isolations. * P

    Article Snippet: Following mounting with ProLong® Gold antifade reagent with 4′,6-diamidino-2-phenylindole (DAPI), images were taken using confocal microscopy (Zeiss LSM 710 NLO) and quantification of the mean intensity was carried out using Adobe Photoshop CS6 software.

    Techniques: In Vitro, Activation Assay, Isolation, Expressing, Real-time Polymerase Chain Reaction, Staining, Western Blot, Marker, Cell Isolation

    The unfolded protein response (UPR) is an early event during in vivo hepatic stellate cell (HSC) activation. a BALB/c mice received a single CCl 4 injection. After the indicated time points, i.e., 2, 6, 10 and 24 h, HSCs were isolated and immediately collected for mRNA analysis of activation and endoplasmic reticulum (ER) stress markers. The graphics show the results of two different HSC isolations (each time point = 3 mice). b Immunofluorescent staining for Bip (green) of HSCs which were isolated, cytospinned and fixed at different time points after a single CCl 4 injection. 4′,6-Diamidino-2-phenylindole (DAPI; blue) was used as a nuclear staining. Graph representing the quantification of % mean intensity in at least 25 cells for each time point, from 3 independent isolations. c mRNA analysis of ER stress markers in HSCs isolated after multiple CCl 4 injections (harvested 24 h after the last injection); n = 5. * P

    Journal: Cell Death & Disease

    Article Title: Unfolded protein response is an early, non-critical event during hepatic stellate cell activation

    doi: 10.1038/s41419-019-1327-5

    Figure Lengend Snippet: The unfolded protein response (UPR) is an early event during in vivo hepatic stellate cell (HSC) activation. a BALB/c mice received a single CCl 4 injection. After the indicated time points, i.e., 2, 6, 10 and 24 h, HSCs were isolated and immediately collected for mRNA analysis of activation and endoplasmic reticulum (ER) stress markers. The graphics show the results of two different HSC isolations (each time point = 3 mice). b Immunofluorescent staining for Bip (green) of HSCs which were isolated, cytospinned and fixed at different time points after a single CCl 4 injection. 4′,6-Diamidino-2-phenylindole (DAPI; blue) was used as a nuclear staining. Graph representing the quantification of % mean intensity in at least 25 cells for each time point, from 3 independent isolations. c mRNA analysis of ER stress markers in HSCs isolated after multiple CCl 4 injections (harvested 24 h after the last injection); n = 5. * P

    Article Snippet: Following mounting with ProLong® Gold antifade reagent with 4′,6-diamidino-2-phenylindole (DAPI), images were taken using confocal microscopy (Zeiss LSM 710 NLO) and quantification of the mean intensity was carried out using Adobe Photoshop CS6 software.

    Techniques: In Vivo, Activation Assay, Mouse Assay, Injection, Isolation, Staining

    c-Jun N-terminal kinase (JNK)-dependent unfolded protein response (UPR) during hepatic stellate cell (HSC) activation. a Freshly isolated mouse HSCs (mHSCs) were treated with the JNK V inhibitor starting at the moment of seeding. Bright-field images of control (days 1 and 7) and JNK inhibitor-treated HSCs (day 7) were taken. b At regular intervals during culture, expression of activation and endoplasmic reticulum (ER) stress markers was determined and compared with untreated cells at the mRNA level by quantitative real-time polymerase chain reaction (qPCR). c Immunofluorescent images for Bip (green, upper left panel) of JNK inhibitor-treated and control cells after 10 h in culture were taken with a confocal microscope. 4′,6-Diamidino-2-phenylindole (DAPI; blue) was used as a nuclear staining. Graph representing the quantification of % mean intensity in at least 25 cells for each time point (right panel). d Control cells (days 1 and 7) and 7-day treated HSCs with JNK inhibitor were fixed and stained for α-smooth muscle actin (α-SMA; red) and DAPI (blue). Images were taken with a confocal microscope. We had to slightly overexpose all groups to be able to observe α-SMA staining in the 10 µM JNK inhibitor-treated group and to keep the same exposure time between the different conditions. Images and graphics are representatives of at least two different HSC isolations. * P

    Journal: Cell Death & Disease

    Article Title: Unfolded protein response is an early, non-critical event during hepatic stellate cell activation

    doi: 10.1038/s41419-019-1327-5

    Figure Lengend Snippet: c-Jun N-terminal kinase (JNK)-dependent unfolded protein response (UPR) during hepatic stellate cell (HSC) activation. a Freshly isolated mouse HSCs (mHSCs) were treated with the JNK V inhibitor starting at the moment of seeding. Bright-field images of control (days 1 and 7) and JNK inhibitor-treated HSCs (day 7) were taken. b At regular intervals during culture, expression of activation and endoplasmic reticulum (ER) stress markers was determined and compared with untreated cells at the mRNA level by quantitative real-time polymerase chain reaction (qPCR). c Immunofluorescent images for Bip (green, upper left panel) of JNK inhibitor-treated and control cells after 10 h in culture were taken with a confocal microscope. 4′,6-Diamidino-2-phenylindole (DAPI; blue) was used as a nuclear staining. Graph representing the quantification of % mean intensity in at least 25 cells for each time point (right panel). d Control cells (days 1 and 7) and 7-day treated HSCs with JNK inhibitor were fixed and stained for α-smooth muscle actin (α-SMA; red) and DAPI (blue). Images were taken with a confocal microscope. We had to slightly overexpose all groups to be able to observe α-SMA staining in the 10 µM JNK inhibitor-treated group and to keep the same exposure time between the different conditions. Images and graphics are representatives of at least two different HSC isolations. * P

    Article Snippet: Following mounting with ProLong® Gold antifade reagent with 4′,6-diamidino-2-phenylindole (DAPI), images were taken using confocal microscopy (Zeiss LSM 710 NLO) and quantification of the mean intensity was carried out using Adobe Photoshop CS6 software.

    Techniques: Activation Assay, Isolation, Expressing, Real-time Polymerase Chain Reaction, Microscopy, Staining

    High-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the fovea for CD 117 (red) and CD44 (green) in the foveola (vertical cross section). ( A ) Immunostaining for CD117 is found predominantly in the interphotoreceptor matrix (white arrowheads). ( B ) Immunostaining for CD44 is found predominantly in the Müller cell apical microvilli in the fovea (unfilled arrowheads) except the foveolar region (white arrow) and in the interphotoreceptor matrix (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the double staining for CD117 and CD44, colocalization yellowish regions, is visible in the interphotoreceptor matrix of the fovea (white arrowheads). Discontinuity of Müller cell apical microvilli is observed in the foveolar region the foveolar region (white arrow).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: High-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the fovea for CD 117 (red) and CD44 (green) in the foveola (vertical cross section). ( A ) Immunostaining for CD117 is found predominantly in the interphotoreceptor matrix (white arrowheads). ( B ) Immunostaining for CD44 is found predominantly in the Müller cell apical microvilli in the fovea (unfilled arrowheads) except the foveolar region (white arrow) and in the interphotoreceptor matrix (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the double staining for CD117 and CD44, colocalization yellowish regions, is visible in the interphotoreceptor matrix of the fovea (white arrowheads). Discontinuity of Müller cell apical microvilli is observed in the foveolar region the foveolar region (white arrow).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Immunostaining, Staining, Double Staining

    High-power-magnification optical microscopy images of a 6-year-old female monkey showing immunostaining of the fovea for GFAP and cellular retinaldehyde-binding protein (CRALBP). ( A ) GFAP (red) expression is visible as intense staining in the inner layer of the foveal Müller cell cone (white arrowheads), the same as is shown in Fig. 1 . ( B ) Immunostaining for CRALBP (green) is slightly observed in the innermost layer of the Müller cell cone (white arrowheads), yet it was negative in most parts of the GFAP-positive area. Moreover, CRALBP staining is visible in the outer plexiform layer around the foveola (unfilled arrowheads). ( C ) DAPI staining (blue). ( D ) In the double-staining for GFAP and CRALBP, almost no colocalization is visible in the inner layer of Müller cell cone of the foveola (white arrowheads).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: High-power-magnification optical microscopy images of a 6-year-old female monkey showing immunostaining of the fovea for GFAP and cellular retinaldehyde-binding protein (CRALBP). ( A ) GFAP (red) expression is visible as intense staining in the inner layer of the foveal Müller cell cone (white arrowheads), the same as is shown in Fig. 1 . ( B ) Immunostaining for CRALBP (green) is slightly observed in the innermost layer of the Müller cell cone (white arrowheads), yet it was negative in most parts of the GFAP-positive area. Moreover, CRALBP staining is visible in the outer plexiform layer around the foveola (unfilled arrowheads). ( C ) DAPI staining (blue). ( D ) In the double-staining for GFAP and CRALBP, almost no colocalization is visible in the inner layer of Müller cell cone of the foveola (white arrowheads).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Immunostaining, Binding Assay, Expressing, Staining, Double Staining

    High-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the foveola (vertical cross section) for nestin (green) and arrestin 4 (red). ( A ) Immunostaining for nestin (green) is mainly observed in the foveolar photoreceptor cell layer (white arrowheads) and the Henle layer (unfilled arrowheads). ( B ) Immunostaining for arrestin 4 (red) is mainly visible in the photoreceptor cell layer alone (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the double-staining for nestin and arrestin 4 including the immunostaining for DAPI (blue), colocalization yellowish regions, is observed in the foveolar photoreceptor layer (white arrowheads).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: High-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the foveola (vertical cross section) for nestin (green) and arrestin 4 (red). ( A ) Immunostaining for nestin (green) is mainly observed in the foveolar photoreceptor cell layer (white arrowheads) and the Henle layer (unfilled arrowheads). ( B ) Immunostaining for arrestin 4 (red) is mainly visible in the photoreceptor cell layer alone (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the double-staining for nestin and arrestin 4 including the immunostaining for DAPI (blue), colocalization yellowish regions, is observed in the foveolar photoreceptor layer (white arrowheads).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Immunostaining, Staining, Double Staining

    Confocal microscopy images of a 4-year-old female monkey showing immunostaining of the foveola (vertical cross section) for GFAP and Tuj-1 and nuclear staining for DAPI (blue). ( A ) Overview section of the foveal pit. Zone 1 indicates B and C . Zone 2 indicates D and E . ( B , C ) GFAP (red) is merged with Tuj-1 (green) in the innermost layer of the foveola (Zone 1, white arrowheads). ( D , E ) No clear evidence of such merging is visible in the area around the foveola (Zone 2, white arrowheads).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: Confocal microscopy images of a 4-year-old female monkey showing immunostaining of the foveola (vertical cross section) for GFAP and Tuj-1 and nuclear staining for DAPI (blue). ( A ) Overview section of the foveal pit. Zone 1 indicates B and C . Zone 2 indicates D and E . ( B , C ) GFAP (red) is merged with Tuj-1 (green) in the innermost layer of the foveola (Zone 1, white arrowheads). ( D , E ) No clear evidence of such merging is visible in the area around the foveola (Zone 2, white arrowheads).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Confocal Microscopy, Immunostaining, Staining

    Medium and high-power-magnification optical microscopy images of a 2-year-old male monkey showing double-immunostaining of the fovea (vertical section) for GFAP and vimentin. ( A ) Medium-power-magnification optical microscopy images. ( B ) Immunostaining for GFAP (red) is observed in the inner layer of the fovea (white arrowheads). ( C ) Vimentin (green) is widely observed from the inner layer to outer layer in the fovea (white arrowheads). ( D ) DAPI staining (blue). ( E ) Weak double-staining for GFAP and vimentin is observed in the surface layer of the foveal slope (unfilled arrowheads).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: Medium and high-power-magnification optical microscopy images of a 2-year-old male monkey showing double-immunostaining of the fovea (vertical section) for GFAP and vimentin. ( A ) Medium-power-magnification optical microscopy images. ( B ) Immunostaining for GFAP (red) is observed in the inner layer of the fovea (white arrowheads). ( C ) Vimentin (green) is widely observed from the inner layer to outer layer in the fovea (white arrowheads). ( D ) DAPI staining (blue). ( E ) Weak double-staining for GFAP and vimentin is observed in the surface layer of the foveal slope (unfilled arrowheads).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Double Immunostaining, Immunostaining, Staining, Double Staining

    Medium-power-magnification optical microscopy images of a 4-year-old female monkey showing double-immunostaining of the foveola (vertical cross section) for GFAP and Tuj-1. ( A ) GFAP (red) expression is visible as intense staining in the inner layer of the foveolar Müller cell cone (white arrowheads), the same as is shown in Fig. 1 . ( B ) Intense immunostaining for Tuj-1 (green) is visible in the inner retinal layer, particularly in the retinal ganglion cell layer (white arrowheads). However, definitive staining is also visible in the innermost layer of the foveal pit (unfilled arrowheads), which spans to the retinal ganglion cell layer. ( C ) DAPI staining (blue). ( D ) Yellowish regions, colocalization of GFAP and Tuj-1,are observed in the surface layer of the foveal pit. (white arrowhead).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: Medium-power-magnification optical microscopy images of a 4-year-old female monkey showing double-immunostaining of the foveola (vertical cross section) for GFAP and Tuj-1. ( A ) GFAP (red) expression is visible as intense staining in the inner layer of the foveolar Müller cell cone (white arrowheads), the same as is shown in Fig. 1 . ( B ) Intense immunostaining for Tuj-1 (green) is visible in the inner retinal layer, particularly in the retinal ganglion cell layer (white arrowheads). However, definitive staining is also visible in the innermost layer of the foveal pit (unfilled arrowheads), which spans to the retinal ganglion cell layer. ( C ) DAPI staining (blue). ( D ) Yellowish regions, colocalization of GFAP and Tuj-1,are observed in the surface layer of the foveal pit. (white arrowhead).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Double Immunostaining, Expressing, Staining, Immunostaining

    Confocal microscopy images of a 6-year-old female monkey showing double-immunostaining of the foveola (vertical cross section) for GFAP (red) and nestin (green), and nuclear staining with DAPI (blue). ( A ) Overview section of foveal pit. Square indicates B and C. ( B , C ) GFAP-positive elongated cells appear to run nearly vertical in the foveola, and some of them, while weakly stained, reach the photoreceptor cell layer (white arrowheads). In the inner layer of the foveola on the same section, the vertically aligned GFAP-positive cells appear to reach the shallow layer of the foveola and are partially merged with nestin. (unfilled arrows). ( D ) Magnified overview of foveal pit. Zone 1 indicates E and Zone 2 indicates F. ( E ) GFAP-positive elongated cells are visible in the foveolar (white arrowhead). ( F ) On the other hand, in the area around the foveola, these GFAP-positive cells are not observed.

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: Confocal microscopy images of a 6-year-old female monkey showing double-immunostaining of the foveola (vertical cross section) for GFAP (red) and nestin (green), and nuclear staining with DAPI (blue). ( A ) Overview section of foveal pit. Square indicates B and C. ( B , C ) GFAP-positive elongated cells appear to run nearly vertical in the foveola, and some of them, while weakly stained, reach the photoreceptor cell layer (white arrowheads). In the inner layer of the foveola on the same section, the vertically aligned GFAP-positive cells appear to reach the shallow layer of the foveola and are partially merged with nestin. (unfilled arrows). ( D ) Magnified overview of foveal pit. Zone 1 indicates E and Zone 2 indicates F. ( E ) GFAP-positive elongated cells are visible in the foveolar (white arrowhead). ( F ) On the other hand, in the area around the foveola, these GFAP-positive cells are not observed.

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Confocal Microscopy, Double Immunostaining, Staining

    High-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the fovea for Ki67. ( A ) Scattered Ki67-positive spots (red) are visible in the retinal innermost layer around the foveola (white arrowheads). Moreover, some parts of the region believed to be the outer plexiform near the inner nuclear layer around the foveola are also Ki67-positive (unfilled arrowheads). ( B ) Immunostaining for Ki67 includes the immunostaining for DAPI (blue).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: High-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the fovea for Ki67. ( A ) Scattered Ki67-positive spots (red) are visible in the retinal innermost layer around the foveola (white arrowheads). Moreover, some parts of the region believed to be the outer plexiform near the inner nuclear layer around the foveola are also Ki67-positive (unfilled arrowheads). ( B ) Immunostaining for Ki67 includes the immunostaining for DAPI (blue).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Immunostaining

    Medium-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the fovea (vertical cross section) for CD 117 (red) and CD44 (green). ( A ) Immunostaining for CD117 is observed predominantly in interphotoreceptor matrix (white arrowheads). ( B ) CD44 is observed predominantly in Müller cell apical microvilli (unfilled arrowheads) in the foveal area and in the interphotoreceptor matrix (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the double-staining for CD117 and CD44, colocalization, a yellowish region, is visible in the interphotoreceptor matrix of the fovea (white arrowheads).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: Medium-power-magnification optical microscopy images of a 4-year-old male monkey showing immunostaining of the fovea (vertical cross section) for CD 117 (red) and CD44 (green). ( A ) Immunostaining for CD117 is observed predominantly in interphotoreceptor matrix (white arrowheads). ( B ) CD44 is observed predominantly in Müller cell apical microvilli (unfilled arrowheads) in the foveal area and in the interphotoreceptor matrix (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the double-staining for CD117 and CD44, colocalization, a yellowish region, is visible in the interphotoreceptor matrix of the fovea (white arrowheads).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Immunostaining, Staining, Double Staining

    Medium-power-magnification optical microscopy images of a 2-year-old male monkey showing immunostaining of the optic disc (vertical cross section) for GFAP and nestin. ( A ) Image showing the region of the optic disc believed to be comprised of astrocytes and the retinal inner layer in the peripapillary area displaying immunostaining for GFAP (red) (white arrowheads). ( B ) The retinal outer layer is stained primarily for nestin (green) (white arrowheads). ( C ) DAPI staining (blue). ( D ) The immunostaining for GFAP is visible in the deep capillary plexus (white arrowheads). The optic disc periphery shows a similar staining pattern to that of the foveolar retina.

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: Medium-power-magnification optical microscopy images of a 2-year-old male monkey showing immunostaining of the optic disc (vertical cross section) for GFAP and nestin. ( A ) Image showing the region of the optic disc believed to be comprised of astrocytes and the retinal inner layer in the peripapillary area displaying immunostaining for GFAP (red) (white arrowheads). ( B ) The retinal outer layer is stained primarily for nestin (green) (white arrowheads). ( C ) DAPI staining (blue). ( D ) The immunostaining for GFAP is visible in the deep capillary plexus (white arrowheads). The optic disc periphery shows a similar staining pattern to that of the foveolar retina.

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Immunostaining, Staining

    High-power-magnification optical microscopy images of a 6-year-old female monkey showing double-immunostaining of the fovea (vertical section) for glial fibrillary acidic protein (GFAP) and nestin. ( A ) Intense immunostaining for GFAP (red) expression is visible in the region where the inner-half of Müller cell cone is believed to be present (white arrowheads), and outer-half of Müller cell cone in the photoreceptor cell layer is less stained. At the boundary between the inner nuclear layer and the outer plexiform layer in the fovea, GFAP-positive cells span to join the GFAP-positive cells in the Müller cell cone (unfilled arrowheads). ( B ) Immunostaining for nestin (green) can be seen mainly in the region where the photoreceptor cell layer (white arrowheads) is located, with the center part of the Müller cell cone displaying weak staining (unfilled arrow). The nestin-positive region spans to the Henle layer (unfilled arrowheads). ( C ) Nuclear staining using 4′, 6-diamidino-2-phenylindole dihydrochloride (DAPI) (blue). ( D ) With double-immunostaining for GFAP and nestin, fluorescence microscopy shows a little overlapping (white arrowheads). At the boundary between the inner nuclear layer and the outer plexiform layer in the fovea, GFAP-positive cells span to join the GFAP-positive cells in the Müller cell cone (unfilled arrowheads).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: High-power-magnification optical microscopy images of a 6-year-old female monkey showing double-immunostaining of the fovea (vertical section) for glial fibrillary acidic protein (GFAP) and nestin. ( A ) Intense immunostaining for GFAP (red) expression is visible in the region where the inner-half of Müller cell cone is believed to be present (white arrowheads), and outer-half of Müller cell cone in the photoreceptor cell layer is less stained. At the boundary between the inner nuclear layer and the outer plexiform layer in the fovea, GFAP-positive cells span to join the GFAP-positive cells in the Müller cell cone (unfilled arrowheads). ( B ) Immunostaining for nestin (green) can be seen mainly in the region where the photoreceptor cell layer (white arrowheads) is located, with the center part of the Müller cell cone displaying weak staining (unfilled arrow). The nestin-positive region spans to the Henle layer (unfilled arrowheads). ( C ) Nuclear staining using 4′, 6-diamidino-2-phenylindole dihydrochloride (DAPI) (blue). ( D ) With double-immunostaining for GFAP and nestin, fluorescence microscopy shows a little overlapping (white arrowheads). At the boundary between the inner nuclear layer and the outer plexiform layer in the fovea, GFAP-positive cells span to join the GFAP-positive cells in the Müller cell cone (unfilled arrowheads).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Double Immunostaining, Immunostaining, Expressing, Staining, Fluorescence

    Medium-power-magnification optical microscopy images of a 2-year-old male monkey showing immunostaining of the retinal extreme periphery (so-called ciliary marginal zone) for GFAP and nestin. ( A ) In the retinal extreme periphery, the retinal inner layer expresses high levels of GFAP (red) (white arrowheads). ( B ) The retinal outer layer expresses high levels of nestin (green) (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the same section, weak double-staining for GFAP and nestin region is visible near the ciliary body, showing a similar staining pattern as the foveolar retina (white arrowheads).

    Journal: Scientific Reports

    Article Title: Immunohistological Study of Monkey Foveal Retina

    doi: 10.1038/s41598-019-41793-y

    Figure Lengend Snippet: Medium-power-magnification optical microscopy images of a 2-year-old male monkey showing immunostaining of the retinal extreme periphery (so-called ciliary marginal zone) for GFAP and nestin. ( A ) In the retinal extreme periphery, the retinal inner layer expresses high levels of GFAP (red) (white arrowheads). ( B ) The retinal outer layer expresses high levels of nestin (green) (white arrowheads). ( C ) DAPI staining (blue). ( D ) In the same section, weak double-staining for GFAP and nestin region is visible near the ciliary body, showing a similar staining pattern as the foveolar retina (white arrowheads).

    Article Snippet: In addition, chemical reaction for DAPI (Dojindo Laboratories, Kumamoto, Japan) and double-immunostaining for GFAP and nestin, GFAP and Tuj-1, nestin and arrestin 4, nestin and vimentin, nestin and neurofilament, CD117 and CD44, and GFAP and CRALBP, respectively, was performed.

    Techniques: Microscopy, Immunostaining, Staining, Double Staining

    Coexpression of p63 with KRT5 and KRT14 in the IPF lung. a , b Sections from IPF human lungs (5 patients) were co-stained either for KRT5 ( green ) and p63 ( red ) ( a left ; b left ), or KRT14 ( red ) and p63 ( green ) ( a right ; b right ) and counterstained with DAPI ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining, for the conducting ( a ) and distal ( b ) healthy airways. c Characteristic KRT5 + pod, stained for KRT5 ( green ) and p63 (red). Arrow: KRT5 + p63 - cell. Full bars = 500 μm. Scattered bars = 100 μm

    Journal: Respiratory Research

    Article Title: Detection and quantification of epithelial progenitor cell populations in human healthy and IPF lungs

    doi: 10.1186/s12931-016-0404-x

    Figure Lengend Snippet: Coexpression of p63 with KRT5 and KRT14 in the IPF lung. a , b Sections from IPF human lungs (5 patients) were co-stained either for KRT5 ( green ) and p63 ( red ) ( a left ; b left ), or KRT14 ( red ) and p63 ( green ) ( a right ; b right ) and counterstained with DAPI ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining, for the conducting ( a ) and distal ( b ) healthy airways. c Characteristic KRT5 + pod, stained for KRT5 ( green ) and p63 (red). Arrow: KRT5 + p63 - cell. Full bars = 500 μm. Scattered bars = 100 μm

    Article Snippet: Nuclei were counterstained with DAPI and the slides mounted in fluorescent mounting medium (Dako, Hamburg, Germany).

    Techniques: Staining

    Costaining of KRT5 with developmental pathway markers in the distal IPF lung. a , b , c , d Sections from IPF human lungs (5 patients) were co-stained for KRT5 ( green ) and ( a ) SHH ( red ), ( b ) GLI1 ( red ), ( c ) SOX9 ( red ), and ( d ) HES1 ( red ), and counterstained with DAPI ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining. Full bars = 100 μm. Scattered bars = 50 μm

    Journal: Respiratory Research

    Article Title: Detection and quantification of epithelial progenitor cell populations in human healthy and IPF lungs

    doi: 10.1186/s12931-016-0404-x

    Figure Lengend Snippet: Costaining of KRT5 with developmental pathway markers in the distal IPF lung. a , b , c , d Sections from IPF human lungs (5 patients) were co-stained for KRT5 ( green ) and ( a ) SHH ( red ), ( b ) GLI1 ( red ), ( c ) SOX9 ( red ), and ( d ) HES1 ( red ), and counterstained with DAPI ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining. Full bars = 100 μm. Scattered bars = 50 μm

    Article Snippet: Nuclei were counterstained with DAPI and the slides mounted in fluorescent mounting medium (Dako, Hamburg, Germany).

    Techniques: Staining

    Proximal-to-distal distribution of KRT5 + and KRT14 + basal cells subpopulations in the healthy human lung. a , b Sections from healthy human lungs (6 donors) were co-stained for Keratin (KRT) 5 ( green ), KRT14 ( red ) and counterstained with DAPI ( blue ). Two representative pictures, with corresponding high magnification panels, are shown for the conducting airways ( a left and right panels ) and the distal airways ( b left and right panels ). Arrow: KRT5 + cell with characteristic luminal epithelial cell morphology. Full bars = 500 μm. Scattered bars = 100 μm. c Isolated primary human bronchial epithelial cells (hBEC) were co-stained for KRT 5 ( green ), KRT14 ( red ) and counterstained with DAPI ( blue ). The panels show: Keratin5-DAPI ( left ), Keratin14-DAPI ( middle ) and merged ( right ) images. Full bars = 500 μm. Scattered bars = 100 μm

    Journal: Respiratory Research

    Article Title: Detection and quantification of epithelial progenitor cell populations in human healthy and IPF lungs

    doi: 10.1186/s12931-016-0404-x

    Figure Lengend Snippet: Proximal-to-distal distribution of KRT5 + and KRT14 + basal cells subpopulations in the healthy human lung. a , b Sections from healthy human lungs (6 donors) were co-stained for Keratin (KRT) 5 ( green ), KRT14 ( red ) and counterstained with DAPI ( blue ). Two representative pictures, with corresponding high magnification panels, are shown for the conducting airways ( a left and right panels ) and the distal airways ( b left and right panels ). Arrow: KRT5 + cell with characteristic luminal epithelial cell morphology. Full bars = 500 μm. Scattered bars = 100 μm. c Isolated primary human bronchial epithelial cells (hBEC) were co-stained for KRT 5 ( green ), KRT14 ( red ) and counterstained with DAPI ( blue ). The panels show: Keratin5-DAPI ( left ), Keratin14-DAPI ( middle ) and merged ( right ) images. Full bars = 500 μm. Scattered bars = 100 μm

    Article Snippet: Nuclei were counterstained with DAPI and the slides mounted in fluorescent mounting medium (Dako, Hamburg, Germany).

    Techniques: Staining, Isolation

    Coexpression of p63 with KRT5 and KRT14 in the healthy human lung. Sections from healthy human lungs (6 donors) were co-stained either for KRT5 ( green ) and p63 ( red ) ( a left ; b left ), or KRT14 ( red ) and p63 ( green ) ( a right ; b right ) and counterstained with DAPI ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining, for the conducting ( a ) and distal ( b ) healthy airways. Arrow: KRT5 - p63 + cell. Full bars = 500 μm. Scattered bars = 100 μm

    Journal: Respiratory Research

    Article Title: Detection and quantification of epithelial progenitor cell populations in human healthy and IPF lungs

    doi: 10.1186/s12931-016-0404-x

    Figure Lengend Snippet: Coexpression of p63 with KRT5 and KRT14 in the healthy human lung. Sections from healthy human lungs (6 donors) were co-stained either for KRT5 ( green ) and p63 ( red ) ( a left ; b left ), or KRT14 ( red ) and p63 ( green ) ( a right ; b right ) and counterstained with DAPI ( blue ). One representative picture, with the corresponding high magnification panel, is shown for each staining, for the conducting ( a ) and distal ( b ) healthy airways. Arrow: KRT5 - p63 + cell. Full bars = 500 μm. Scattered bars = 100 μm

    Article Snippet: Nuclei were counterstained with DAPI and the slides mounted in fluorescent mounting medium (Dako, Hamburg, Germany).

    Techniques: Staining

    KRT5 + and KRT14 + basal cell subpopulations in the IPF lung. Sections from IPF human lungs (5 patients) were co-stained for KRT5 (green), KRT14 (red) and counterstained with DAPI ( blue ). a Two representative pictures, with corresponding high magnification panels, are shown for the conducting airways. b 4 representative pictures, with corresponding high magnification panels, illustrate the distribution patterns (metaplastic, simple and pod-like) of KRT5 + and KRT14 + basal cell subpopulations in the distal IPF lung. Full bars = 500 μm. Scattered bars = 100 μm

    Journal: Respiratory Research

    Article Title: Detection and quantification of epithelial progenitor cell populations in human healthy and IPF lungs

    doi: 10.1186/s12931-016-0404-x

    Figure Lengend Snippet: KRT5 + and KRT14 + basal cell subpopulations in the IPF lung. Sections from IPF human lungs (5 patients) were co-stained for KRT5 (green), KRT14 (red) and counterstained with DAPI ( blue ). a Two representative pictures, with corresponding high magnification panels, are shown for the conducting airways. b 4 representative pictures, with corresponding high magnification panels, illustrate the distribution patterns (metaplastic, simple and pod-like) of KRT5 + and KRT14 + basal cell subpopulations in the distal IPF lung. Full bars = 500 μm. Scattered bars = 100 μm

    Article Snippet: Nuclei were counterstained with DAPI and the slides mounted in fluorescent mounting medium (Dako, Hamburg, Germany).

    Techniques: Staining