r26 tdt  (Worthington Biochemical)


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    Worthington Biochemical r26 tdt
    Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × <t>R26-TdT</t> mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).
    R26 Tdt, supplied by Worthington Biochemical, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/r26 tdt/product/Worthington Biochemical
    Average 92 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    r26 tdt - by Bioz Stars, 2020-07
    92/100 stars

    Images

    1) Product Images from "Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart"

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart

    Journal: Circulation

    doi: 10.1161/CIRCULATIONAHA.118.035210

    Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × R26-TdT mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).
    Figure Legend Snippet: Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × R26-TdT mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).

    Techniques Used: Mouse Assay, Immunohistochemistry, Quantitation Assay, Flow Cytometry, Cytometry, Fluorescence, Staining

    Cardiac Sca-1 + Cells Contribute to the Vasculature Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using constitutive Sca-1 (gene name Ly6a) Cre gene-targeted mice. B. Representative confocal micrographs of histological sections from hearts of Ly6a +/Cre × R26-TdT mice at either postnatal day 1, 1.5 months of age, or 3 months of age, showing continual expansion of TdTomato + cells (red) in the heart. Scale bars = 100 μm. C, D. Immunohistochemistry on cardiac histological sections was performed to detect CD31 (white) along with endogenous TdTomato fluorescence (red). DAPI (blue) was used to visualize nuclei. TdTomato + endothelial cells were seen in small clusters at postnatal day 1 ( C ) and throughout the heart at 1.5 months of age ( D ). Scale bars = 10 μm. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from dissociated Ly6a +/Cre × R26-TdT mouse hearts at 3 months of age ( n =3) analyzed using antibodies against Sca-1 and CD31. The first plot shows Sca-1 positivity by fluorochrome-conjugated antibody staining (Sca-1), versus CD31 positivity also by antibody (CD31). The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 − gate (lower right quadrant) shown in the first plot as indicated. The final plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 + gate (upper right quadrant) shown in the first plot as indicated.
    Figure Legend Snippet: Cardiac Sca-1 + Cells Contribute to the Vasculature Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using constitutive Sca-1 (gene name Ly6a) Cre gene-targeted mice. B. Representative confocal micrographs of histological sections from hearts of Ly6a +/Cre × R26-TdT mice at either postnatal day 1, 1.5 months of age, or 3 months of age, showing continual expansion of TdTomato + cells (red) in the heart. Scale bars = 100 μm. C, D. Immunohistochemistry on cardiac histological sections was performed to detect CD31 (white) along with endogenous TdTomato fluorescence (red). DAPI (blue) was used to visualize nuclei. TdTomato + endothelial cells were seen in small clusters at postnatal day 1 ( C ) and throughout the heart at 1.5 months of age ( D ). Scale bars = 10 μm. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from dissociated Ly6a +/Cre × R26-TdT mouse hearts at 3 months of age ( n =3) analyzed using antibodies against Sca-1 and CD31. The first plot shows Sca-1 positivity by fluorochrome-conjugated antibody staining (Sca-1), versus CD31 positivity also by antibody (CD31). The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 − gate (lower right quadrant) shown in the first plot as indicated. The final plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 + gate (upper right quadrant) shown in the first plot as indicated.

    Techniques Used: Mouse Assay, Immunohistochemistry, Fluorescence, Flow Cytometry, Cytometry, Quantitation Assay, Staining

    2) Product Images from "Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart"

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart

    Journal: Circulation

    doi: 10.1161/CIRCULATIONAHA.118.035210

    Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × R26-TdT mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).
    Figure Legend Snippet: Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × R26-TdT mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).

    Techniques Used: Mouse Assay, Immunohistochemistry, Quantitation Assay, Flow Cytometry, Cytometry, Fluorescence, Staining

    Cardiac Sca-1 + Cells Contribute to the Vasculature Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using constitutive Sca-1 (gene name Ly6a) Cre gene-targeted mice. B. Representative confocal micrographs of histological sections from hearts of Ly6a +/Cre × R26-TdT mice at either postnatal day 1, 1.5 months of age, or 3 months of age, showing continual expansion of TdTomato + cells (red) in the heart. Scale bars = 100 μm. C, D. Immunohistochemistry on cardiac histological sections was performed to detect CD31 (white) along with endogenous TdTomato fluorescence (red). DAPI (blue) was used to visualize nuclei. TdTomato + endothelial cells were seen in small clusters at postnatal day 1 ( C ) and throughout the heart at 1.5 months of age ( D ). Scale bars = 10 μm. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from dissociated Ly6a +/Cre × R26-TdT mouse hearts at 3 months of age ( n =3) analyzed using antibodies against Sca-1 and CD31. The first plot shows Sca-1 positivity by fluorochrome-conjugated antibody staining (Sca-1), versus CD31 positivity also by antibody (CD31). The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 − gate (lower right quadrant) shown in the first plot as indicated. The final plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 + gate (upper right quadrant) shown in the first plot as indicated.
    Figure Legend Snippet: Cardiac Sca-1 + Cells Contribute to the Vasculature Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using constitutive Sca-1 (gene name Ly6a) Cre gene-targeted mice. B. Representative confocal micrographs of histological sections from hearts of Ly6a +/Cre × R26-TdT mice at either postnatal day 1, 1.5 months of age, or 3 months of age, showing continual expansion of TdTomato + cells (red) in the heart. Scale bars = 100 μm. C, D. Immunohistochemistry on cardiac histological sections was performed to detect CD31 (white) along with endogenous TdTomato fluorescence (red). DAPI (blue) was used to visualize nuclei. TdTomato + endothelial cells were seen in small clusters at postnatal day 1 ( C ) and throughout the heart at 1.5 months of age ( D ). Scale bars = 10 μm. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from dissociated Ly6a +/Cre × R26-TdT mouse hearts at 3 months of age ( n =3) analyzed using antibodies against Sca-1 and CD31. The first plot shows Sca-1 positivity by fluorochrome-conjugated antibody staining (Sca-1), versus CD31 positivity also by antibody (CD31). The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 − gate (lower right quadrant) shown in the first plot as indicated. The final plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 + gate (upper right quadrant) shown in the first plot as indicated.

    Techniques Used: Mouse Assay, Immunohistochemistry, Fluorescence, Flow Cytometry, Cytometry, Quantitation Assay, Staining

    Related Articles

    Flow Cytometry:

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart
    Article Snippet: .. For flow cytometry analysis of dissociated cardiac non-myocytes from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , or Ly6a +/MerCreMer × R26-eGFP mice, single-cell suspensions were prepared via repeated rounds of enzymatic digestion and trituration using collagenase type IV (Worthington # ) and dispase II (Roche #10165859001) according to “Protocol #2” from Pinto et. al. . .. For flow cytometry analysis of whole bone marrow from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , Ly 6a +/+ , or Ly6a +/MerCreMer mice, cells were isolated by cannulating dissected tibias and femurs with a 25-gauge needle and flushing with 5–10 mL of flow cytometry staining buffer as previously described .

    Cytometry:

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart
    Article Snippet: .. For flow cytometry analysis of dissociated cardiac non-myocytes from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , or Ly6a +/MerCreMer × R26-eGFP mice, single-cell suspensions were prepared via repeated rounds of enzymatic digestion and trituration using collagenase type IV (Worthington # ) and dispase II (Roche #10165859001) according to “Protocol #2” from Pinto et. al. . .. For flow cytometry analysis of whole bone marrow from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , Ly 6a +/+ , or Ly6a +/MerCreMer mice, cells were isolated by cannulating dissected tibias and femurs with a 25-gauge needle and flushing with 5–10 mL of flow cytometry staining buffer as previously described .

    Mouse Assay:

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart
    Article Snippet: .. For flow cytometry analysis of dissociated cardiac non-myocytes from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , or Ly6a +/MerCreMer × R26-eGFP mice, single-cell suspensions were prepared via repeated rounds of enzymatic digestion and trituration using collagenase type IV (Worthington # ) and dispase II (Roche #10165859001) according to “Protocol #2” from Pinto et. al. . .. For flow cytometry analysis of whole bone marrow from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , Ly 6a +/+ , or Ly6a +/MerCreMer mice, cells were isolated by cannulating dissected tibias and femurs with a 25-gauge needle and flushing with 5–10 mL of flow cytometry staining buffer as previously described .

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    Worthington Biochemical r26 tdt
    Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × <t>R26-TdT</t> mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).
    R26 Tdt, supplied by Worthington Biochemical, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/r26 tdt/product/Worthington Biochemical
    Average 92 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
    r26 tdt - by Bioz Stars, 2020-07
    92/100 stars
      Buy from Supplier

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    Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × R26-TdT mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).

    Journal: Circulation

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart

    doi: 10.1161/CIRCULATIONAHA.118.035210

    Figure Lengend Snippet: Cardiac Sca-1 + Cells Contribute Few Cardiomyocytes Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using Ly6a -Cre gene-targeted mice. B. Immunohistochemistry was performed on cardiac histological sections from Ly6a +/Cre × R26-TdT mice at 3 months of age using antibodies against sarcomeric α-actinin (white) and PCM1 (purple). DAPI (blue) was used to visualize nuclei. Representative confocal micrograph shows a rare TdTomato + cardiomyocyte (yellow box). Scale bar: 100 μm. C. High-magnification confocal micrographs of the yellow boxed area denoted in B . Yellow arrowhead indicates a TdTomato + cardiomyocyte (α-actinin + PCM1 + ). Scale bars = 10 μm. D. Quantitation of TdTomato + cardiomyocytes from histological sections of hearts from Ly6a +/Cre × R26-TdT mice at 3 months of age. Quantitation is shown from hearts of n =5 mice over 144 histological sections and 303260 total cardiomyocytes counted. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from the bone marrow of Ly6a +/Cre × R26-TdT mice at 3 months of age ( n =3). The first plot shows forward (FSC-A) versus side (SSC-A) scatter to determine size distribution of the bone marrow. The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter from the gate shown in the first plot as indicated ( E ). Cells within this second gate were stained with antibodies against Sca-1 and CD45, which showed primarily mature CD45 + leukocytes that were TdTomato + ( F ).

    Article Snippet: For flow cytometry analysis of dissociated cardiac non-myocytes from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , or Ly6a +/MerCreMer × R26-eGFP mice, single-cell suspensions were prepared via repeated rounds of enzymatic digestion and trituration using collagenase type IV (Worthington # ) and dispase II (Roche #10165859001) according to “Protocol #2” from Pinto et. al. .

    Techniques: Mouse Assay, Immunohistochemistry, Quantitation Assay, Flow Cytometry, Cytometry, Fluorescence, Staining

    Cardiac Sca-1 + Cells Contribute to the Vasculature Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using constitutive Sca-1 (gene name Ly6a) Cre gene-targeted mice. B. Representative confocal micrographs of histological sections from hearts of Ly6a +/Cre × R26-TdT mice at either postnatal day 1, 1.5 months of age, or 3 months of age, showing continual expansion of TdTomato + cells (red) in the heart. Scale bars = 100 μm. C, D. Immunohistochemistry on cardiac histological sections was performed to detect CD31 (white) along with endogenous TdTomato fluorescence (red). DAPI (blue) was used to visualize nuclei. TdTomato + endothelial cells were seen in small clusters at postnatal day 1 ( C ) and throughout the heart at 1.5 months of age ( D ). Scale bars = 10 μm. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from dissociated Ly6a +/Cre × R26-TdT mouse hearts at 3 months of age ( n =3) analyzed using antibodies against Sca-1 and CD31. The first plot shows Sca-1 positivity by fluorochrome-conjugated antibody staining (Sca-1), versus CD31 positivity also by antibody (CD31). The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 − gate (lower right quadrant) shown in the first plot as indicated. The final plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 + gate (upper right quadrant) shown in the first plot as indicated.

    Journal: Circulation

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart

    doi: 10.1161/CIRCULATIONAHA.118.035210

    Figure Lengend Snippet: Cardiac Sca-1 + Cells Contribute to the Vasculature Throughout Postnatal Growth. A. Experimental scheme and timeline for genetic lineage tracing studies in this figure using constitutive Sca-1 (gene name Ly6a) Cre gene-targeted mice. B. Representative confocal micrographs of histological sections from hearts of Ly6a +/Cre × R26-TdT mice at either postnatal day 1, 1.5 months of age, or 3 months of age, showing continual expansion of TdTomato + cells (red) in the heart. Scale bars = 100 μm. C, D. Immunohistochemistry on cardiac histological sections was performed to detect CD31 (white) along with endogenous TdTomato fluorescence (red). DAPI (blue) was used to visualize nuclei. TdTomato + endothelial cells were seen in small clusters at postnatal day 1 ( C ) and throughout the heart at 1.5 months of age ( D ). Scale bars = 10 μm. E, F. Representative flow cytometry plots ( E ) and quantitation ( F ) from dissociated Ly6a +/Cre × R26-TdT mouse hearts at 3 months of age ( n =3) analyzed using antibodies against Sca-1 and CD31. The first plot shows Sca-1 positivity by fluorochrome-conjugated antibody staining (Sca-1), versus CD31 positivity also by antibody (CD31). The second plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 − gate (lower right quadrant) shown in the first plot as indicated. The final plot shows TdTomato + cells (endogenous TdTomato fluorescence) versus side scatter within the Sca-1 + CD31 + gate (upper right quadrant) shown in the first plot as indicated.

    Article Snippet: For flow cytometry analysis of dissociated cardiac non-myocytes from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , or Ly6a +/MerCreMer × R26-eGFP mice, single-cell suspensions were prepared via repeated rounds of enzymatic digestion and trituration using collagenase type IV (Worthington # ) and dispase II (Roche #10165859001) according to “Protocol #2” from Pinto et. al. .

    Techniques: Mouse Assay, Immunohistochemistry, Fluorescence, Flow Cytometry, Cytometry, Quantitation Assay, Staining

    Inducible Genetic Lineage Tracing in the Adult Mouse Tracks Sca-1 + Cells During Aging. A. Experimental scheme for genetic lineage tracing studies in this figure using a tamoxifen-inducible Ly6a -MerCreMer gene-targeted mouse line and the R26-eGFP reporter line. B. eGFP reporter expression is dependent on tamoxifen induction, as untreated Ly6a +/MerCreMer × R26-eGFP mice aged for one year do not show eGFP + cells across multiple tissues surveyed. C. Ly6a +/MerCreMer × R26-eGFP mice treated with tamoxifen starting at adulthood (3 mos) out to 1 yr of age showed endothelial cell labeling (CD31, red) of eGFP + cells (green) throughout the heart, lung, liver, intestine, and kidney. Yellow boxes indicate areas shown to the right of each image in an enlarged view. Other cell types previously reported to express Sca-1 are also labeled with eGFP. DAPI (blue) was used to visualize nuclei. Scale bars = 100 μm.

    Journal: Circulation

    Article Title: Genetic Lineage Tracing of Sca-1+ Cells Reveals Endothelial but Not Myogenic Contribution to the Murine Heart

    doi: 10.1161/CIRCULATIONAHA.118.035210

    Figure Lengend Snippet: Inducible Genetic Lineage Tracing in the Adult Mouse Tracks Sca-1 + Cells During Aging. A. Experimental scheme for genetic lineage tracing studies in this figure using a tamoxifen-inducible Ly6a -MerCreMer gene-targeted mouse line and the R26-eGFP reporter line. B. eGFP reporter expression is dependent on tamoxifen induction, as untreated Ly6a +/MerCreMer × R26-eGFP mice aged for one year do not show eGFP + cells across multiple tissues surveyed. C. Ly6a +/MerCreMer × R26-eGFP mice treated with tamoxifen starting at adulthood (3 mos) out to 1 yr of age showed endothelial cell labeling (CD31, red) of eGFP + cells (green) throughout the heart, lung, liver, intestine, and kidney. Yellow boxes indicate areas shown to the right of each image in an enlarged view. Other cell types previously reported to express Sca-1 are also labeled with eGFP. DAPI (blue) was used to visualize nuclei. Scale bars = 100 μm.

    Article Snippet: For flow cytometry analysis of dissociated cardiac non-myocytes from Ly6a +/+ × R26-TdT , Ly6a +/Cre × R26-TdT , or Ly6a +/MerCreMer × R26-eGFP mice, single-cell suspensions were prepared via repeated rounds of enzymatic digestion and trituration using collagenase type IV (Worthington # ) and dispase II (Roche #10165859001) according to “Protocol #2” from Pinto et. al. .

    Techniques: Expressing, Mouse Assay, Labeling