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 2351 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    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 "Cytosolic Phospholipase A2α Promotes Pulmonary Inflammation and Systemic Disease during Streptococcus pneumoniae Infection"

    Article Title: Cytosolic Phospholipase A2α Promotes Pulmonary Inflammation and Systemic Disease during Streptococcus pneumoniae Infection

    Journal: Infection and Immunity

    doi: 10.1128/IAI.00280-17

    S. pneumoniae infection of airway epithelial cells triggers arachidonic acid (AA) release. (A) H292 cell monolayers were labeled with [ 3 H]AA as detailed in Materials and Methods and infected with the indicated S. pneumoniae ( S.p. ) strains (each of a different capsular serotype) or with B. subtilis strain 168 at a multiplicity of infection (MOI) of 10 (1 ×10 7 CFU/monolayer). Supernatants were collected at the indicated times postinfection, and radioactive counts released into the supernatants were measured by scintillation counting. Labeled H292 cell monolayers treated with PMA and HBSS+Ca/Mg were used as positive and negative controls, respectively. Shown are results from a representative of two experiments. (B) H292 cell monolayers labeled with [ 3 H]AA were treated with the indicated concentrations of pan-PLA 2 inhibitors ACA or ONO-RS-082 or the DAG lipase inhibitor RHC-80267 (labeled as RHC) prior to infection with S. pneumoniae TIGR4, as detailed in Materials and Methods. Labeled H292 cell monolayers treated with HBSS+Ca/Mg were used as negative controls. Radioactive counts released into supernatants were determined by scintillation counting. Shown are results from a representative of three experiments. *, P
    Figure Legend Snippet: S. pneumoniae infection of airway epithelial cells triggers arachidonic acid (AA) release. (A) H292 cell monolayers were labeled with [ 3 H]AA as detailed in Materials and Methods and infected with the indicated S. pneumoniae ( S.p. ) strains (each of a different capsular serotype) or with B. subtilis strain 168 at a multiplicity of infection (MOI) of 10 (1 ×10 7 CFU/monolayer). Supernatants were collected at the indicated times postinfection, and radioactive counts released into the supernatants were measured by scintillation counting. Labeled H292 cell monolayers treated with PMA and HBSS+Ca/Mg were used as positive and negative controls, respectively. Shown are results from a representative of two experiments. (B) H292 cell monolayers labeled with [ 3 H]AA were treated with the indicated concentrations of pan-PLA 2 inhibitors ACA or ONO-RS-082 or the DAG lipase inhibitor RHC-80267 (labeled as RHC) prior to infection with S. pneumoniae TIGR4, as detailed in Materials and Methods. Labeled H292 cell monolayers treated with HBSS+Ca/Mg were used as negative controls. Radioactive counts released into supernatants were determined by scintillation counting. Shown are results from a representative of three experiments. *, P

    Techniques Used: Infection, Labeling, Proximity Ligation Assay

    Ablation of cPLA 2 α leads to decreased bacteremia and increased survival in an otherwise lethal S. pneumoniae lung challenge. cPLA 2 α −/− mice or their littermate WT controls were intratracheally inoculated with TIGR4 (see Materials and Methods). A control group of WT mice received PBS. (A) Bacteremia was determined by plating tail vein blood at the specified time points. Shown are results from a combination of two experiments. Death of all infected WT mice is indicated by a dagger. (B) Survival of animals was monitored over a 7-day postinfection period. The log rank (Mantel-Cox) test was performed for survival curve analysis. The experiment was performed twice, with similar results. Shown are results from a combination of two experiments. A total of 8 mice were used per group; panels A and B represent the same cohorts of mice.
    Figure Legend Snippet: Ablation of cPLA 2 α leads to decreased bacteremia and increased survival in an otherwise lethal S. pneumoniae lung challenge. cPLA 2 α −/− mice or their littermate WT controls were intratracheally inoculated with TIGR4 (see Materials and Methods). A control group of WT mice received PBS. (A) Bacteremia was determined by plating tail vein blood at the specified time points. Shown are results from a combination of two experiments. Death of all infected WT mice is indicated by a dagger. (B) Survival of animals was monitored over a 7-day postinfection period. The log rank (Mantel-Cox) test was performed for survival curve analysis. The experiment was performed twice, with similar results. Shown are results from a combination of two experiments. A total of 8 mice were used per group; panels A and B represent the same cohorts of mice.

    Techniques Used: Mouse Assay, Infection

    3) Product Images from "Inflammatory Chemokine Transport and Presentation in HEV"

    Article Title: Inflammatory Chemokine Transport and Presentation in HEV

    Journal: The Journal of Experimental Medicine

    doi:

    Short-term homing of adoptively transferred L-selectin + CCR2 + monocytes to inflamed PLNs is mediated by MCP-1. (A) Flow cytometry of PBMCs from CX3CR1 +/GFP knockin mice stained for the monocyte-macrophage markers F4/80 or CD11b (Mac-1) reveals two populations of GFP + leukocytes based upon differential expression of GFP, L-selectin (CD62L), and CCR2. The F4/80 + GFP − cells were identified as eosinophils by their high side scatter (data not shown). (B) Flow cytometric analysis of leukocyte populations in homing assays. CX3CR1 +/GFP PBMCs were injected intravenously into wild-type recipients 5–7 d after induction of skin inflammation by CFA/KLH injection. 4 h later, GFP + donor populations in recipient blood and inflamed PLNs were compared with input PBMCs. To avoid counting of contaminating GFP + NK cells, all samples were stained with anti-NK1.1. Numbers in dot plots indicate the percentage of GFP + cells in the GFP MED and GFP HIGH gates in one representative experiment (out of 6). (C) GFP MED cells (white bars), but not GFP HIGH cells (black bars) were preferentially recruited to inflamed PLNs. Data presented as mean ± SEM, n = 6 mice per group. ** P
    Figure Legend Snippet: Short-term homing of adoptively transferred L-selectin + CCR2 + monocytes to inflamed PLNs is mediated by MCP-1. (A) Flow cytometry of PBMCs from CX3CR1 +/GFP knockin mice stained for the monocyte-macrophage markers F4/80 or CD11b (Mac-1) reveals two populations of GFP + leukocytes based upon differential expression of GFP, L-selectin (CD62L), and CCR2. The F4/80 + GFP − cells were identified as eosinophils by their high side scatter (data not shown). (B) Flow cytometric analysis of leukocyte populations in homing assays. CX3CR1 +/GFP PBMCs were injected intravenously into wild-type recipients 5–7 d after induction of skin inflammation by CFA/KLH injection. 4 h later, GFP + donor populations in recipient blood and inflamed PLNs were compared with input PBMCs. To avoid counting of contaminating GFP + NK cells, all samples were stained with anti-NK1.1. Numbers in dot plots indicate the percentage of GFP + cells in the GFP MED and GFP HIGH gates in one representative experiment (out of 6). (C) GFP MED cells (white bars), but not GFP HIGH cells (black bars) were preferentially recruited to inflamed PLNs. Data presented as mean ± SEM, n = 6 mice per group. ** P

    Techniques Used: Flow Cytometry, Cytometry, Knock-In, Mouse Assay, Staining, Expressing, Injection

    Rapid, sustained induction of MCP-1 in inflamed skin precipitates monocyte/macrophage accumulation in draining PLNs. (A) Time course of monocyte/macrophage recruitment to PLNs of wild-type and MCP-1 −/ − mice. CD11b + F4/80 + leukocytes were counted in subiliac PLNs at different times after intracutaneous injection of CFA/KLH into the ipsilateral flank. Monocyte numbers in the contralateral, noninflamed subiliac PLNs are shown for comparison. † P
    Figure Legend Snippet: Rapid, sustained induction of MCP-1 in inflamed skin precipitates monocyte/macrophage accumulation in draining PLNs. (A) Time course of monocyte/macrophage recruitment to PLNs of wild-type and MCP-1 −/ − mice. CD11b + F4/80 + leukocytes were counted in subiliac PLNs at different times after intracutaneous injection of CFA/KLH into the ipsilateral flank. Monocyte numbers in the contralateral, noninflamed subiliac PLNs are shown for comparison. † P

    Techniques Used: Mouse Assay, Injection

    4) 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

    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.
    Figure Legend 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.

    Techniques Used: Expressing, Mouse Assay, Labeling

    5) Product Images from "Expression of neuroimmune semaphorins 4A and 4D and their receptors in the lung is enhanced by allergen and vascular endothelial growth factor"

    Article Title: Expression of neuroimmune semaphorins 4A and 4D and their receptors in the lung is enhanced by allergen and vascular endothelial growth factor

    Journal: BMC Immunology

    doi: 10.1186/1471-2172-12-30

    Regulation of lung CD72 expression by allergen and VEGF . Sema4D receptor CD72 expression in mouse lung tissue and cells was analyzed by immunohistochemistry and flow cytometry (A-B). (A) Formalin-fixed paraffin-embedded lung tissue sections obtained from either control or OVA-treated WT mice were deparaffinized and immunohistochemistry was performed using appropriate Abs as described in Materials and Methods. Frozen lung tissue sections were used for VEGF tg mice. CD72 is the most abundantly expressed molecule in the lungs of PBS-treated mice. Its expression is further upregulated with OVA treatment. Note that subsets of inflammatory granulocytes and lymphocytes express CD72 (red arrow). In VEGF tg lungs CD72 expression was mainly targeted to inflammatory macrophages (red arrow). Red arrowhead points to the lung epithelial cell CD72 expression. (B) CD72 is expressed on lung MHCII+ cells including DC and is upregulated by OVA. Both CD4+ and CD8+ T cells in the lung express CD72 at low level which is further upregulated by allergen on CD4+ T cells.
    Figure Legend Snippet: Regulation of lung CD72 expression by allergen and VEGF . Sema4D receptor CD72 expression in mouse lung tissue and cells was analyzed by immunohistochemistry and flow cytometry (A-B). (A) Formalin-fixed paraffin-embedded lung tissue sections obtained from either control or OVA-treated WT mice were deparaffinized and immunohistochemistry was performed using appropriate Abs as described in Materials and Methods. Frozen lung tissue sections were used for VEGF tg mice. CD72 is the most abundantly expressed molecule in the lungs of PBS-treated mice. Its expression is further upregulated with OVA treatment. Note that subsets of inflammatory granulocytes and lymphocytes express CD72 (red arrow). In VEGF tg lungs CD72 expression was mainly targeted to inflammatory macrophages (red arrow). Red arrowhead points to the lung epithelial cell CD72 expression. (B) CD72 is expressed on lung MHCII+ cells including DC and is upregulated by OVA. Both CD4+ and CD8+ T cells in the lung express CD72 at low level which is further upregulated by allergen on CD4+ T cells.

    Techniques Used: Expressing, Immunohistochemistry, Flow Cytometry, Cytometry, Formalin-fixed Paraffin-Embedded, Mouse Assay

    Regulation of lung Tim-2 expression by allergen and VEGF . Immunohistochemical (A-B) and flow cytometric (C) detection of Tim-2 expression in lung (A) and lymphoid (B) tissues, and on lung T cells (C). (A-B) Note positive Tim-2 staining on different lung cells besides lymphocytes in allergen- or VEGF-exposed mouse lungs. Red arrows point to Tim-2+ APC-like cells and granulocyte. Inserts show high magnification fields (100x) with marker-positive cells (lymphocytes). (C) Flow cytometry analysis of cells from lung enzymatic digests obtained from PBS- and allergen-treated mice showed an increase in lung Tim-2+ T cells in OVA-treated mice. The histograms show the percentage of Tim-2+CD3+ cells in the lung (clear histogram) as compared to the appropriate isotype control stained CD3+ cells (gray histogram).
    Figure Legend Snippet: Regulation of lung Tim-2 expression by allergen and VEGF . Immunohistochemical (A-B) and flow cytometric (C) detection of Tim-2 expression in lung (A) and lymphoid (B) tissues, and on lung T cells (C). (A-B) Note positive Tim-2 staining on different lung cells besides lymphocytes in allergen- or VEGF-exposed mouse lungs. Red arrows point to Tim-2+ APC-like cells and granulocyte. Inserts show high magnification fields (100x) with marker-positive cells (lymphocytes). (C) Flow cytometry analysis of cells from lung enzymatic digests obtained from PBS- and allergen-treated mice showed an increase in lung Tim-2+ T cells in OVA-treated mice. The histograms show the percentage of Tim-2+CD3+ cells in the lung (clear histogram) as compared to the appropriate isotype control stained CD3+ cells (gray histogram).

    Techniques Used: Expressing, Immunohistochemistry, Flow Cytometry, Staining, Marker, Cytometry, Mouse Assay

    6) 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

    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.
    Figure Legend 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.

    Techniques Used: Expressing, Mouse Assay, Labeling

    7) 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

    8) 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

    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.
    Figure Legend 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.

    Techniques Used: Expressing, Mouse Assay, Labeling

    9) Product Images from "Inflammatory Chemokine Transport and Presentation in HEV"

    Article Title: Inflammatory Chemokine Transport and Presentation in HEV

    Journal: The Journal of Experimental Medicine

    doi:

    Short-term homing of adoptively transferred L-selectin + CCR2 + monocytes to inflamed PLNs is mediated by MCP-1. (A) Flow cytometry of PBMCs from CX3CR1 +/GFP knockin mice stained for the monocyte-macrophage markers F4/80 or CD11b (Mac-1) reveals two populations of GFP + leukocytes based upon differential expression of GFP, L-selectin (CD62L), and CCR2. The F4/80 + GFP − cells were identified as eosinophils by their high side scatter (data not shown). (B) Flow cytometric analysis of leukocyte populations in homing assays. CX3CR1 +/GFP PBMCs were injected intravenously into wild-type recipients 5–7 d after induction of skin inflammation by CFA/KLH injection. 4 h later, GFP + donor populations in recipient blood and inflamed PLNs were compared with input PBMCs. To avoid counting of contaminating GFP + NK cells, all samples were stained with anti-NK1.1. Numbers in dot plots indicate the percentage of GFP + cells in the GFP MED and GFP HIGH gates in one representative experiment (out of 6). (C) GFP MED cells (white bars), but not GFP HIGH cells (black bars) were preferentially recruited to inflamed PLNs. Data presented as mean ± SEM, n = 6 mice per group. ** P
    Figure Legend Snippet: Short-term homing of adoptively transferred L-selectin + CCR2 + monocytes to inflamed PLNs is mediated by MCP-1. (A) Flow cytometry of PBMCs from CX3CR1 +/GFP knockin mice stained for the monocyte-macrophage markers F4/80 or CD11b (Mac-1) reveals two populations of GFP + leukocytes based upon differential expression of GFP, L-selectin (CD62L), and CCR2. The F4/80 + GFP − cells were identified as eosinophils by their high side scatter (data not shown). (B) Flow cytometric analysis of leukocyte populations in homing assays. CX3CR1 +/GFP PBMCs were injected intravenously into wild-type recipients 5–7 d after induction of skin inflammation by CFA/KLH injection. 4 h later, GFP + donor populations in recipient blood and inflamed PLNs were compared with input PBMCs. To avoid counting of contaminating GFP + NK cells, all samples were stained with anti-NK1.1. Numbers in dot plots indicate the percentage of GFP + cells in the GFP MED and GFP HIGH gates in one representative experiment (out of 6). (C) GFP MED cells (white bars), but not GFP HIGH cells (black bars) were preferentially recruited to inflamed PLNs. Data presented as mean ± SEM, n = 6 mice per group. ** P

    Techniques Used: Flow Cytometry, Cytometry, Knock-In, Mouse Assay, Staining, Expressing, Injection

    Rapid, sustained induction of MCP-1 in inflamed skin precipitates monocyte/macrophage accumulation in draining PLNs. (A) Time course of monocyte/macrophage recruitment to PLNs of wild-type and MCP-1 −/ − mice. CD11b + F4/80 + leukocytes were counted in subiliac PLNs at different times after intracutaneous injection of CFA/KLH into the ipsilateral flank. Monocyte numbers in the contralateral, noninflamed subiliac PLNs are shown for comparison. † P
    Figure Legend Snippet: Rapid, sustained induction of MCP-1 in inflamed skin precipitates monocyte/macrophage accumulation in draining PLNs. (A) Time course of monocyte/macrophage recruitment to PLNs of wild-type and MCP-1 −/ − mice. CD11b + F4/80 + leukocytes were counted in subiliac PLNs at different times after intracutaneous injection of CFA/KLH into the ipsilateral flank. Monocyte numbers in the contralateral, noninflamed subiliac PLNs are shown for comparison. † P

    Techniques Used: Mouse Assay, Injection

    10) Product Images from "Depletion of CD11c+ cells in the CD11c.DTR model drives expansion of unique CD64+ Ly6C+ monocytes that are poised to release TNF-α"

    Article Title: Depletion of CD11c+ cells in the CD11c.DTR model drives expansion of unique CD64+ Ly6C+ monocytes that are poised to release TNF-α

    Journal: European Journal of Immunology

    doi: 10.1002/eji.201545789

    CCR2-independent expansion of DT-Ly6C + monocytes in vivo. (A) CD11c.DTR.Ccr2 −/− mice or CD11c.DTR controls were injected with PBS ( n = 3–4) or DT ( n = 2–4) and the expansion of monocytes analyzed in the spleen 48 h later. Graph shows the number of CD11b + Ly6C + cells ± SEM, PBS versus DT in CD11c.DTR.Ccr2 −/− mice p = 0.029. Data are pooled from three independent experiments. (B) Schematic diagram describing the competitive chimera experiments. (C) Left: representative flow cytometry plots showing CD11b + Ly6C + monocytes in pregated bulk splenic CD45.2 + hematopoietic cells from PBS- or DT-treated chimeras, and the percentage of WT (CD45.1 + CD45.2 + ) or Ccr2 −/− (CD45.2 + ) cells in the monocyte gate. Right: graph shows the frequency of WT or Ccr2 −/− CD11b + Ly6C + cells in the spleens of PBS- or DT-treated chimeras; PBS-treated ( n = 6) and DT-treated ( n = 6) mice. WT cells PBS versus DT p = 0.0260, Ccr2 −/− cells p = 0.0411. Data are pooled from two independent experiments. Statistical analyses were carried out with a Mann–Whitney U test.
    Figure Legend Snippet: CCR2-independent expansion of DT-Ly6C + monocytes in vivo. (A) CD11c.DTR.Ccr2 −/− mice or CD11c.DTR controls were injected with PBS ( n = 3–4) or DT ( n = 2–4) and the expansion of monocytes analyzed in the spleen 48 h later. Graph shows the number of CD11b + Ly6C + cells ± SEM, PBS versus DT in CD11c.DTR.Ccr2 −/− mice p = 0.029. Data are pooled from three independent experiments. (B) Schematic diagram describing the competitive chimera experiments. (C) Left: representative flow cytometry plots showing CD11b + Ly6C + monocytes in pregated bulk splenic CD45.2 + hematopoietic cells from PBS- or DT-treated chimeras, and the percentage of WT (CD45.1 + CD45.2 + ) or Ccr2 −/− (CD45.2 + ) cells in the monocyte gate. Right: graph shows the frequency of WT or Ccr2 −/− CD11b + Ly6C + cells in the spleens of PBS- or DT-treated chimeras; PBS-treated ( n = 6) and DT-treated ( n = 6) mice. WT cells PBS versus DT p = 0.0260, Ccr2 −/− cells p = 0.0411. Data are pooled from two independent experiments. Statistical analyses were carried out with a Mann–Whitney U test.

    Techniques Used: In Vivo, Mouse Assay, Injection, Flow Cytometry, Cytometry, MANN-WHITNEY

    11) 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

    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.
    Figure Legend 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.

    Techniques Used: Expressing, Mouse Assay, Labeling

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    Worthington Biochemical chong ly
    Spectroscopic analysis of cis or trans <t>-3-chloroallyl-Lys-4</t> <t>H3-21</t> ( 3 and 4 ) treated GST-LSD1. UV/Vis spectra of native LSD1 (blue) shows the distinctive two maxima of the fully oxidized flavin and the one electron reduced semiquinone. A) LSD1 treated with
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    Spectroscopic analysis of cis or trans -3-chloroallyl-Lys-4 H3-21 ( 3 and 4 ) treated GST-LSD1. UV/Vis spectra of native LSD1 (blue) shows the distinctive two maxima of the fully oxidized flavin and the one electron reduced semiquinone. A) LSD1 treated with

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Spectroscopic analysis of cis or trans -3-chloroallyl-Lys-4 H3-21 ( 3 and 4 ) treated GST-LSD1. UV/Vis spectra of native LSD1 (blue) shows the distinctive two maxima of the fully oxidized flavin and the one electron reduced semiquinone. A) LSD1 treated with

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques:

    Spectroscopic and MALDI-TOF analysis of hydrazino-Lys-4 H3-21 ( 18 ) treated GST-LSD1. A) UV/Vis spectra of native LSD1 (blue) shows the distinctive two maxima of the fully oxidized flavin and the one electron reduced semiquinone. LSD1 treated with 18 (red)

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Spectroscopic and MALDI-TOF analysis of hydrazino-Lys-4 H3-21 ( 18 ) treated GST-LSD1. A) UV/Vis spectra of native LSD1 (blue) shows the distinctive two maxima of the fully oxidized flavin and the one electron reduced semiquinone. LSD1 treated with 18 (red)

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques:

    Synthesis of peptide inhibitors cis -3-chloroallyl-Lys-4 H3-21 ( 3 ), trans -3-chloroallyl-Lys-4 H3-21 ( 4 ) and hydrazino-Lys-4 H3-21 ( 18 ). Incorporation of Fmoc-hydroxynorleucine in the four position of H3-21 yields the resin bound peptide. Mesylation of

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Synthesis of peptide inhibitors cis -3-chloroallyl-Lys-4 H3-21 ( 3 ), trans -3-chloroallyl-Lys-4 H3-21 ( 4 ) and hydrazino-Lys-4 H3-21 ( 18 ). Incorporation of Fmoc-hydroxynorleucine in the four position of H3-21 yields the resin bound peptide. Mesylation of

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques:

    Time- and concentration-dependent inactivation of GST-LSD1 by cis -3-chloroallyl-Lys-4 H3-21 ( 3 ) and trans -3-chloroallyl-Lys-4 H3-21 ( 4 ). A and C) Steady-state progress curves obtained for the inactivation of LSD1 in the presence of concentrations of

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Time- and concentration-dependent inactivation of GST-LSD1 by cis -3-chloroallyl-Lys-4 H3-21 ( 3 ) and trans -3-chloroallyl-Lys-4 H3-21 ( 4 ). A and C) Steady-state progress curves obtained for the inactivation of LSD1 in the presence of concentrations of

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques: Concentration Assay

    Inhibition of GST-LSD1 by endo -cyclopropyl-Lys-4 H3-21 ( 6 ) and endo -dimethylcyclopropyl-Lys-4 H3-21 ( 7 ). A and C) Steady-state progress curves obtained for the inhibition of LSD1 in the presence of concentrations of 6 ranging from 0 to 49.6 µM

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Inhibition of GST-LSD1 by endo -cyclopropyl-Lys-4 H3-21 ( 6 ) and endo -dimethylcyclopropyl-Lys-4 H3-21 ( 7 ). A and C) Steady-state progress curves obtained for the inhibition of LSD1 in the presence of concentrations of 6 ranging from 0 to 49.6 µM

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques: Inhibition

    Proposed mechanism of inactivation of GST-LSD1 by hydrazino-Lys-4 H3-21 ( 18 ). Pathway a , two electron oxidation of 18 by the flavin yields the diazene. A second oxidation to the primary diazonium species produces a powerful electrophilic site at the alpha

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Proposed mechanism of inactivation of GST-LSD1 by hydrazino-Lys-4 H3-21 ( 18 ). Pathway a , two electron oxidation of 18 by the flavin yields the diazene. A second oxidation to the primary diazonium species produces a powerful electrophilic site at the alpha

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques:

    Time- and concentration-dependent inactivation of GST-LSD1 by hydrazino-Lys-4 H3-21 ( 18 ). A) Steady-state progress curves obtained for the inactivation of LSD1 in the presence of concentrations of 18 ranging from 0.1 to 0.4 µM. B) Rate constants

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Time- and concentration-dependent inactivation of GST-LSD1 by hydrazino-Lys-4 H3-21 ( 18 ). A) Steady-state progress curves obtained for the inactivation of LSD1 in the presence of concentrations of 18 ranging from 0.1 to 0.4 µM. B) Rate constants

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques: Concentration Assay

    Proposed mechanism of inactivation of GST-LSD1 by cis -3-chloroallyl-Lys-4 H3-21 ( 3 ). Flavin mediated two electron oxidation of 3 yields an α,β-unsaturated iminium species. Pathway a ) Nucleophilic attack by the N 5 of the reduced flavin

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: Proposed mechanism of inactivation of GST-LSD1 by cis -3-chloroallyl-Lys-4 H3-21 ( 3 ). Flavin mediated two electron oxidation of 3 yields an α,β-unsaturated iminium species. Pathway a ) Nucleophilic attack by the N 5 of the reduced flavin

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques:

    MALDI-TOF assay demonstrating the inactivation of GST-LSD1 by phenelzine. A) LSD1 preincubated with phenelzine prior to exposure to dimethyl-Lys-4 H3-21 substrate fails to appreciably demethylate. B) LSD1 efficiently demethylates dimethyl-Lys-4 H3-21

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: MALDI-TOF assay demonstrating the inactivation of GST-LSD1 by phenelzine. A) LSD1 preincubated with phenelzine prior to exposure to dimethyl-Lys-4 H3-21 substrate fails to appreciably demethylate. B) LSD1 efficiently demethylates dimethyl-Lys-4 H3-21

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques:

    MALDI-TOF analysis of GST-LSD1 incubated with cis or trans -3-chloroallyl-Lys-4 H3-21 ( 3 and 4 ). A) Inactivator 3 is seen as a minor peak, while a new signal corresponding to the predicted mass of a 3 -FAD adduct is now apparent. Additionally, a signal

    Journal: Journal of the American Chemical Society

    Article Title: Comparative Analysis of Small Molecules and Histone Substrate Analogs as LSD1 Lysine Demethylase Inhibitors

    doi: 10.1021/ja909996p

    Figure Lengend Snippet: MALDI-TOF analysis of GST-LSD1 incubated with cis or trans -3-chloroallyl-Lys-4 H3-21 ( 3 and 4 ). A) Inactivator 3 is seen as a minor peak, while a new signal corresponding to the predicted mass of a 3 -FAD adduct is now apparent. Additionally, a signal

    Article Snippet: The 150 µL reactions were initiated by the addition of 50 µl of buffered substrate (dimethyl-Lys-4 H3-21) solution to reaction mixtures (100 µl) consisting of 50 mM HEPES buffer (pH 7.5), 0.1 mM 4-aminoantipyrine, 1 mM 3,5-dichloro-2-hydroxybenzenesulfonic acid, 0.76 µM horseradish peroxidase (Worthington Biochemical Corporation), and 185 nM GST-LSD1.

    Techniques: Incubation