Washless double staining of unfixed nuclei for flow cytometric analysis of DNA and a nuclear antigen (Ki‐67 or bromodeoxyuridine)

Larsen, Jørgen K.; Christensen, Ib Jarle; Christiansen, Jens; Mortensen, Børge Thing

doi:10.1002/cyto.990120508

Cited by 35 publications

(18 citation statements)

References 20 publications

(14 reference statements)

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“…Next, the orthotopic and metastatic xenograft tumor cells in mice that formed metastatic lesions (xenograft #4) were sorted by FACS, and the cell cycle of the cancer cells was analyzed by an anti‐Ki‐67 mAb and propidium iodide (PI) staining (Larsen et al . ) (Fig. C, F).…”

Section: Resultsmentioning

confidence: 80%

Absence of primary cilia in cell cycle‐arrested human breast cancer cells

et al. 2013

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Previous studies using cultured cells showed that primary cilia are present in quiescent cells, but are absent in proliferating cells. We studied here the relationship between the presence or absence of primary cilia and the cell cycle arrest of normal epithelial cells and cancer cells in the human normal breast and breast cancer tissues. In normal breast tissues, although most epithelial cells were nonproliferating as estimated by the immunofluorescence staining of the proliferation marker Ki-67, primary cilia were present only in 20-40% of the epithelial cells. In breast cancer tissues, primary cilia were not observed in any of the breast cancer cells. Furthermore, primary cilia were hardly observed in the nonproliferating cancer cells in the orthotopic and metastatic human breast cancer xenograft tumors in mice. These results indicate that the absence of primary cilia does not necessarily represent the proliferating phases of normal epithelial cells and cancer cells.

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Section: Resultsmentioning

confidence: 80%

Absence of primary cilia in cell cycle‐arrested human breast cancer cells

et al. 2013

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“…Also, cell-cycle-dependent antigen expression, e.g., S-phase specific PCNA (13), Ki-67 (14), and cyclins (9,10), can be studied more accurately when cells are also stained for DNA. Therefore, we conclude that the addition of TP3 eliminates the spectral cross talk problems caused by PI and improves the analysis of antigen expression in heterogeneous cell populations in combination with DNA-ploidy measurements.…”

Section: Discussionmentioning

confidence: 99%

Improved single laser measurement of two cellular antigens and DNA-ploidy by the combined use of propidium iodide and TO-PRO-3 iodide

1997

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“…The cells of the dissociated PDX stained with Alexa Fluor 488-conjugated mouse anti-human HLA-A, B, C mAb and the cell line-derived tumor cells expressing ZsGreen1 were analyzed and sorted using FACS Aria I (BD Biosciences). In the cell cycle analysis, the sorted cells were subsequently permeabilized by a detergent solution (0.5% Triton-X 100 in PBS, 0.5 mM EDTA, pH 7.2), and then stained with propidium iodide (Sigma) with addition of RNase (Sigma) as previously described [21]. In the ex vivo culture experiment, the high expression level of cell surface CXCR4 was determined as the signal level that includes more than 50% of the orthotopic cancer cells and less than 5% of the metastasized cancer cells in the lung on average.…”

Section: Methodsmentioning

confidence: 99%

Downregulation of CXCR4 in Metastasized Breast Cancer Cells and Implication in Their Dormancy

et al. 2015

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Our understanding of the mechanism of cancer dormancy is emerging, but the underlying mechanisms are not fully understood. Here we analyzed mouse xenograft tumors derived from human breast cancer tissue and the human breast cancer cell line MDA-MB-231 to identify the molecules associated with cancer dormancy. In immunohistological examination using the proliferation marker Ki-67, the tumors included both proliferating and dormant cancer cells, but the number of dormant cells was remarkably increased when they metastasized to the lung. In the gene expression analysis of the orthotopic cancer cells by a single-cell multiplex real-time quantitative reverse transcription PCR followed by flow cytometric analysis, restrained cellular proliferation was associated with downregulation of the chemokine receptor CXCR4. In the immunohistological and flow cytometric analyses, the expression level of CXCR4 in the metastasized cancer cells was decreased compared with that in the cancer cells in orthotopic tumors, although the expression level of the CXCR4 ligand CXCL12 was not reduced in the lung. In addition, the proliferation of the metastasized cancer cells was further decreased by the CXCR4 antagonist administration. In the ex vivo culture of the metastasized cancer cells, the expression level of CXCR4 was increased, and in the xenotransplantation of ex vivo cultured cancer cells, the expression level of CXCR4 was again decreased in the metastasized cancer cells in the lung. These findings indicate that CXCR4 is downregulated in metastasized breast cancer cells and implicated in their dormancy.

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Washless double staining of unfixed nuclei for flow cytometric analysis of DNA and a nuclear antigen (Ki‐67 or bromodeoxyuridine)

Cited by 35 publications

References 20 publications

Absence of primary cilia in cell cycle‐arrested human breast cancer cells

Absence of primary cilia in cell cycle‐arrested human breast cancer cells

Improved single laser measurement of two cellular antigens and DNA-ploidy by the combined use of propidium iodide and TO-PRO-3 iodide

Downregulation of CXCR4 in Metastasized Breast Cancer Cells and Implication in Their Dormancy

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