Immunofluorescence analysis of cytokeratin 8/18 staining is a sensitive assay for the detection of cell apoptosis

Dong,Qiao‑Mei; Ling,Chun; Zhao,Li

doi:10.3892/ol.2015.2856

Immunofluorescence analysis of cytokeratin 8/18 staining is a sensitive assay for the detection of cell apoptosis

Authors:
- Qiao‑Mei Dong
- Chun Ling
- Li Zhao
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Affiliations: Central Laboratory, The First Hospital of Lanzhou University, Lanzhou, Gansu 730000, P.R. China
Published online on: January 7, 2015 https://doi.org/10.3892/ol.2015.2856
Pages: 1227-1230

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Abstract

Apoptosis is one of the major types of programmed cell death. During this process, cells experience a series of morphological and biochemical changes. Flow cytometric analysis of Annexin V staining of cell surface phosphatidylserine, in combination with a DNA‑staining dye to probe the permeability of the cell membrane, is an established method for detecting apoptosis. The present study aimed to show that the immunofluorescence analysis of cytokeratin (CK) 8/18 staining may provide a new and sensitive assay for the detection of apoptotic cells. Tumor cells were treated with 20 µM cisplatin to induce apoptosis. Following 12 and 24 h of cisplatin treatment, cells were collected and stained with 4',6‑diamidine‑2'‑phenylindole dihydrochloride (DAPI) and fluorescein‑labeled anti‑CK8/18 antibody. The apoptotic cells were subsequently examined by fluorescence microscopy. Annexin V‑fluorescein isothiocyanate/propidium iodide staining followed by flow cytometric analysis confirmed that cisplatin was able to induce apoptosis in tumor cells. Immunofluorescence analysis demonstrated that apoptotic cells had a distinct CK8/18 staining pattern. In living cells, CK8/18 was uniformly distributed in the cytoplasm and cytosol; however in the apoptotic cells with a condensed and/or fragmented apoptotic nucleus (as identified by DAPI staining), fluorescein‑labeled anti‑CK8/18 antibody exhibited unusual punctate and/or bubbly staining in the cytosol. In the apoptotic cells that could not be identified by DAPI staining, fluorescein‑labeled CK8/18 displayed polarized aggregated staining in the cytosol. These results indicate that fluorescein‑conjugated CK8/18 may be a useful and sensitive indicator of cell apoptosis.

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