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Optimized Ki-67 staining in murine cells: a tool to determine cell proliferation

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Abstract

The reliable analysis of the cell cycle status has become increasingly relevant for scientific and clinical work, especially for the determination of tumor cell growth. One established method to characterize the proliferation activity of cells is the analysis of the Ki-67 protein. Ki-67 is expressed in the nucleus during the whole cell cycle except for the G0 phase. Several different protocols exist for the examination of the Ki-67 protein in tissue and cell culture, but most of them are defined for human cells. For the analysis of the Ki-67 protein in murine tissue and cell culture there is a variety of protocols existing which recommend different fixation and permeabilization reagents or special kits. In this study, we established a reliable protocol for Ki-67 staining in murine cells and tissue based on PFA fixation, which can be used not only for flow cytometry but also for immunofluorescence microscopy analysis. We tested our protocol successfully with three different Ki-67 anti-mouse antibodies in cell culture, regenerating liver tissue and mouse melanoma tumor to demonstrate the general applicability.

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Acknowledgements

The authors acknowledge the excellent support of the flow-cytometry core facility at the medical faculty of Bonn University. Furthermore, they thank Chrystel Flores for perfect technical assistance. C. W. was supported by the Else Kröner-Forschungskolleg Bonn and Bonfor in Bonn; C. K., Z. A., I. P. and A. H. are members of the excellence cluster ‘‘ImmunoSensation’’ at Bonn University.

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Author notes

  1. C. Graefe and L. Eichhorn are equally contributing.

Authors and Affiliations

  1. Department of Anaesthesiology and Intensive Care Medicine, University Hospital Bonn, Bonn, Germany

    C. Graefe, L. Eichhorn, J. Kleiner, A. Hoeft, S. Frede & C. K. Weisheit

  2. Flow Cytometry Core Facility, Institute of Experimental Immunology, University Hospital Bonn, Bonn, Germany

    P. Wurst

  3. Institute of Experimental Immunology, University Hospital Bonn, Bonn, Germany

    P. Wurst, I. Panetas, Z. Abdulla & C. Kurts

  4. Department of Oncology, Hematology and Rheumatology, University Hospital Bonn, Bonn, Germany

    A. Heine

  5. Koordinationsstelle Research Infrastructure, University Hospital Bonn, Bonn, Germany

    E. Endl

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  1. C. Graefe
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  2. L. Eichhorn
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  3. P. Wurst
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  4. J. Kleiner
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  11. E. Endl
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Contributions

Conceived and designed the experiments: CW, LE, EE, ZA. Performed the experiments: CG, LE, JK, PW, AH, IP. Analyzed the data: CG, LE, CW, EE, AH. Wrote the paper: CG, CW, SF, AH, CK, EE.

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Correspondence to C. K. Weisheit.

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Graefe, C., Eichhorn, L., Wurst, P. et al. Optimized Ki-67 staining in murine cells: a tool to determine cell proliferation. Mol Biol Rep 46, 4631–4643 (2019). https://doi.org/10.1007/s11033-019-04851-2

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  • DOI: https://doi.org/10.1007/s11033-019-04851-2

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