Cell Synchronization Techniques to Study the Action of CDK Inhibitors

  • Beatriz Pérez-Benavente
  • Rosa FarràsEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1336)


Cell synchronization techniques have been used for the studies of mechanisms involved in cell cycle regulation. Synchronization involves the enrichment of subpopulations of cells in specific stages of the cell cycle. These subpopulations are then used to study regulatory mechanisms of the cell cycle such as DNA synthesis, gene expression, protein synthesis, protein phosphorylation, protein degradation, and development of new drugs (e.g., CDK inhibitors). Here, we describe several protocols for synchronization of cells from different phases of the cell cycle. We also describe protocols for determining cell viability and mitotic index and for validating the synchrony of the cells by flow cytometry.

Key words

Cell cycle DNA content Flow cytometry Mitotic index Synchronization 


  1. 1.
    Morgan DO (2007) The cell cycle: principles of control, Primers in biology. New Science Press Ltd in association with Oxford University Press, LondonGoogle Scholar
  2. 2.
    Banfalvi G (2011) Overview of cell synchronization. Methods Mol Biol 761:1–23PubMedCrossRefGoogle Scholar
  3. 3.
    Rosner M, Schipany K, Hengstschlager M (2013) Merging high-quality biochemical fractionation with a refined flow cytometry approach to monitor nucleocytoplasmic protein expression throughout the unperturbed mammalian cell cycle. Nat Protoc 8(3):602–626PubMedCrossRefGoogle Scholar
  4. 4.
    Campisi J, Morreo G, Pardee AB (1984) Kinetics of G1 transit following brief starvation for serum factors. Exp Cell Res 152(2):459–466PubMedCrossRefGoogle Scholar
  5. 5.
    Masters JR (2002) HeLa cells 50 years on: the good, the bad and the ugly. Nat Rev Cancer 2(4):315–319PubMedCrossRefGoogle Scholar
  6. 6.
    Farras R, Baldin V, Gallach S, Acquaviva C, Bossis G, Jariel-Encontre I, Piechaczyk M (2008) JunB breakdown in mid-/late G2 is required for down-regulation of cyclin A2 levels and proper mitosis. Mol Cell Biol 28(12):4173–4187. MCB.01620-07 [pii]PubMedCentralPubMedCrossRefGoogle Scholar
  7. 7.
    Stein GS, Borun TW (1972) The synthesis of acidic chromosomal proteins during the cell cycle of HeLa S-3 cells. I. The accelerated accumulation of acidic residual nuclear protein before the initiation of DNA replication. J Cell Biol 52(2):292–307PubMedCentralPubMedCrossRefGoogle Scholar
  8. 8.
    Harper JV (2005) Synchronization of cell populations in G1/S and G2/M phases of the cell cycle. Methods Mol Biol 296:157–166PubMedGoogle Scholar
  9. 9.
    Heintz N, Sive HL, Roeder RG (1983) Regulation of human histone gene expression: kinetics of accumulation and changes in the rate of synthesis and in the half-lives of individual histone mRNAs during the HeLa cell cycle. Mol Cell Biol 3(4):539–550PubMedCentralPubMedGoogle Scholar
  10. 10.
    Pines J, Hunter T (1989) Isolation of a human cyclin cDNA: evidence for cyclin mRNA and protein regulation in the cell cycle and for interaction with p34cdc2. Cell 58(5):833–846, 0092-8674(89)90936-7 [pii]PubMedCrossRefGoogle Scholar
  11. 11.
    Kohn KW, Jackman J, O’Connor PM (1994) Cell cycle control and cancer chemotherapy. J Cell Biochem 54(4):440–452PubMedCrossRefGoogle Scholar
  12. 12.
    Perez-Benavente B, Garcia JL, Rodriguez MS, Pineda-Lucena A, Piechaczyk M, Font de Mora J, Farras R (2013) GSK3-SCF(FBXW7) targets JunB for degradation in G2 to preserve chromatid cohesion before anaphase. Oncogene 32(17):2189–2199PubMedCrossRefGoogle Scholar
  13. 13.
    Ferrero M, Ferragud J, Orlando L, Valero L, Sanchez del Pino M, Farras R, Font de Mora J (2011) Phosphorylation of AIB1 at mitosis is regulated by CDK1/CYCLIN B. PLoS One 6(12), e28602PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Xu B, Kim S, Kastan MB (2001) Involvement of Brca1 in S-phase and G(2)-phase checkpoints after ionizing irradiation. Mol Cell Biol 21(10):3445–3450PubMedCentralPubMedCrossRefGoogle Scholar
  15. 15.
    Elvin P, Evans CW (1983) Cell adhesiveness and the cell cycle: correlation in synchronized Balb/c 3T3 cells. Biol Cell 48(1):1–9PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2016

Authors and Affiliations

  1. 1.Oncogenic Signalling LaboratoryCentro de Investigación Príncipe Felipe de ValenciaValenciaSpain
  2. 2.Biochemistry and Molecular Biology DepartmentUniversidad de ValenciaValenciaSpain

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