The aim of this protocol is to provide a comprehensive description of the materials, equipment and reproducible methods to detect and analyze anaphase bridges in immunofluorescence microscopy using DAPI to detect cells that failed to completely segregate during mitosis. It describes the process of cell preparation, staining and microscopic settings for detection of anaphase bridges. The protocol has been adapted from our previous publication (Aschacher et al., 2016).
[Background] During cell division it is vital for the maintenance of genome integrity that the genetic material is fully separated. For various reasons this process can be dysfunctional and as a result the sister chromatids are connected by DNA bridges, which most frequently happens during anaphase. Especially chromosomal fragile sites are associated with anaphase bridges (e.g., unprotected and unstable telomeres). Breakage, deletion, translocation non-disjunction and changes in chromosome number at these sites are often linked with cancer and other genetic diseases. Two types of anaphase bridges are described, the ultrafine DNA bridges, that cannot be detected by DAPI staining and the chromatin bridges, which are visualized by DAPI (Germann et al., 2014). The latter is described subsequently.
This protocol describes a fast and simple method for the detection and calculation of anaphase bridges to provide an additional assay for telomere attrition in any publications.
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