IMAGE: The cover for issue 24 of Oncotarget features Figure 1, “Workflow for enumeration of cleaved caspase-3 bleb?positive [CC3(bleb)+] cells in FFPE canine tumor tissue, ” from Dull, et al…. view more
Existing microscopy-based methods of detecting apoptosis, such as TUNEL (terminal deoxynucleotidyl transferase dUTP nick-end labeling), have limited quantitative capabilities due to insufficient signal-to-noise ratios. Researchers at the National Cancer Institute and National Cancer Institute-Frederick have addressed this issue via development of a highly specific apoptosis assay designed for immunofluorescence microscopy analysis of fixed core needle biopsy specimens.
The high specificity of this novel assay is due to the measurement of colocalization of two independent markers of apoptosis. The first marker is γH2AX, the serine 139-phosphorylated form of histone H2AX that is found at sites of DNA double-strand breaks and can arise either as a consequence of DNA damage or due to the DNA laddering that occurs during apoptosis. The second marker is the formation of cleaved caspase-3 aggregates or puncta, which arise concurrently with apoptosis-associated plasma membrane blebbing.
Cleaved caspase-3 (CC3) is used as a marker of apoptosis in sandwich ELISA and immunohistochemical analyses, but detection of this activated caspase in many non-apoptotic cells has
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