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The global steps required for CQ-FISH. After proper tissue embedding and staining according to the protocol, all samples are imaged on the same day to avoid signal variation with time. Shown are representative images from CQ-FISH analysis, immunostained for cardiomyocyte-specific marker cardiac troponin T (cTnT; green), telomeric probe (red dots), and DAPI for nuclei (blue). Scale bar, 10 μm. White dashed outlines mark the areas used for telomere analysis within the nucleus of each cardiomyocyte. Unspecific fluorescence (yellow arrowheads) outside the nucleus is not included in telomere analysis. The <t>Telometer</t> software is used to manually identify the nuclear area of the cell type of interest. Telomeres are automatically identified within the encircled nuclear area and analyzed, and parameters are measured for graphing.
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The global steps required for CQ-FISH. After proper tissue embedding and staining according to the protocol, all samples are imaged on the same day to avoid signal variation with time. Shown are representative images from CQ-FISH analysis, immunostained for cardiomyocyte-specific marker cardiac troponin T (cTnT; green), telomeric probe (red dots), and DAPI for nuclei (blue). Scale bar, 10 μm. White dashed outlines mark the areas used for telomere analysis within the nucleus of each cardiomyocyte. Unspecific fluorescence (yellow arrowheads) outside the nucleus is not included in telomere analysis. The <t>Telometer</t> software is used to manually identify the nuclear area of the cell type of interest. Telomeres are automatically identified within the encircled nuclear area and analyzed, and parameters are measured for graphing.
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The global steps required for CQ-FISH. After proper tissue embedding and staining according to the protocol, all samples are imaged on the same day to avoid signal variation with time. Shown are representative images from CQ-FISH analysis, immunostained for cardiomyocyte-specific marker cardiac troponin T (cTnT; green), telomeric probe (red dots), and DAPI for nuclei (blue). Scale bar, 10 μm. White dashed outlines mark the areas used for telomere analysis within the nucleus of each cardiomyocyte. Unspecific fluorescence (yellow arrowheads) outside the nucleus is not included in telomere analysis. The Telometer software is used to manually identify the nuclear area of the cell type of interest. Telomeres are automatically identified within the encircled nuclear area and analyzed, and parameters are measured for graphing.

Journal: Nature protocols

Article Title: Evaluation of telomere length in human cardiac tissues using cardiac quantitative FISH

doi: 10.1038/nprot.2017.082

Figure Lengend Snippet: The global steps required for CQ-FISH. After proper tissue embedding and staining according to the protocol, all samples are imaged on the same day to avoid signal variation with time. Shown are representative images from CQ-FISH analysis, immunostained for cardiomyocyte-specific marker cardiac troponin T (cTnT; green), telomeric probe (red dots), and DAPI for nuclei (blue). Scale bar, 10 μm. White dashed outlines mark the areas used for telomere analysis within the nucleus of each cardiomyocyte. Unspecific fluorescence (yellow arrowheads) outside the nucleus is not included in telomere analysis. The Telometer software is used to manually identify the nuclear area of the cell type of interest. Telomeres are automatically identified within the encircled nuclear area and analyzed, and parameters are measured for graphing.

Article Snippet: A convenient method is to use the custom software module Telometer, developed at the Johns Hopkins School of Medicine (the program and documentation are freely available for download at http://demarzolab.pathology.jhmi.edu/telometer/index.html ) 9 , 19 .

Techniques: Staining, Marker, Fluorescence, Software