未处理和经阿非迪霉素处理的DT40细胞的甲醛固定中期染色体的免疫染色
Immunostaining of Formaldehyde-fixed Metaphase Chromosome from Untreated and Aphidicolin-treated DT40 Cells.
作者信息
Oestergaard Vibe H
机构信息
Department of Biology, University of Copenhagen, Ole Maaloees Vej 5, Copenhagen N, Denmark.
出版信息
Bio Protoc. 2017 May 5;7(9):e2259. doi: 10.21769/BioProtoc.2259.
During mitosis chromosomes are condensed into dense X-shaped structures that allow for microscopic determination of karyotype as well as inspection of chromosome morphology. This protocol describes a method to perform immunostaining of formaldehyde-fixed metaphase chromosomes from the avian cell line DT40. It was developed to characterize the localization of YFP-tagged TopBP1 on mitotic chromosomes and specifically determine the percentage of TopBP1 foci that formed on breaks/gaps as well as ends of individual metaphase macrochromosomes ( Pedersen , 2015 ). For this purpose immunostaining of YFP was applied. However, the protocol may be optimized for other cell lines or epitopes.
在有丝分裂期间，染色体浓缩成致密的X形结构，这使得显微镜下确定核型以及检查染色体形态成为可能。本方案描述了一种对来自禽类细胞系DT40的甲醛固定中期染色体进行免疫染色的方法。该方法是为了表征黄色荧光蛋白（YFP）标记的TopBP1在有丝分裂染色体上的定位，并具体确定在单个中期大染色体的断裂/间隙以及末端形成的TopBP1焦点的百分比而开发的（佩德森，2015年）。为此应用了YFP免疫染色。然而，该方案可针对其他细胞系或表位进行优化。
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