Skip to main content
SpringerLink
Log in

Automated time-lapse microscopy and high-resolution tracking of cell migration

  • Original Paper
  • Published:
Cytotechnology Aims and scope Submit manuscript

Abstract

We describe a novel fully automated high-throughput time-lapse microscopy system and evaluate its performance for precisely tracking the motility of several glioma and osteoblastic cell lines. Use of this system revealed cell motility behavior not discernable with conventional techniques by collecting data (1) from closely spaced time points (minutes), (2) over long periods (hours to days), (3) from multiple areas of interest, (4) in parallel under several different experimental conditions. Quantitation of true individual and average cell velocity and path length was obtained with high spatial and temporal resolution in “scratch” or “wound healing” assays. This revealed unique motility dynamics of drug-treated and adhesion molecule-transfected cells and, thus, this is a considerable improvement over current methods of measurement and analysis. Several fluorescent vital labeling methods commonly used for end-point analyses (GFP expression, DiO lipophilic dye, and Qtracker nanocrystals) were found to be useful for time-lapse studies under specific conditions that are described. To illustrate one application, fluorescently labeled tumor cells were seeded onto cell monolayers expressing ectopic adhesion molecules, and this resulted in consistently reduced tumor cell migration velocities. These highly quantitative time-lapse analysis methods will promote the creation of new cell motility assays and increase the resolution and accuracy of existing assays.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Besson A, Gurian-West M, Schmidt A, Hall A, Roberts JM (2004) p27Kip1 modulates cell migration through the regulation of RhoA activation. Genes Dev 18:862–876

    Article  CAS  Google Scholar 

  • Cretu A, Fotos JS, Little BW, Galileo DS (2005) Human and rat glioma growth, invasion, and vascularization in a novel chick embryo brain tumor model. Clin Exper Metas 22:225–236

    Article  Google Scholar 

  • Edme N, Downward J, Thiery JP, Boyer B (2002) Ras induces NBT-II epithelial cell scattering through the coordinate activities of Rac and MAPK pathways. J Cell Sci 115:2591–2601

    CAS  Google Scholar 

  • Endo Y, Wolf V, Muraiso K, Kamijo K, Soon L, Uren A, Barshishat-Kupper M, Rubin JS (2005) Wnt-3a-dependent cell motility involves RhoA activation and is specifically regulated by dishevelled-2. J Biol Chem 280:777–786

    CAS  Google Scholar 

  • Galileo DS, Gee AP, Linser PJ (1991) Neurons are replenished in cultures of embryonic chick optic tectum after immunomagnetic depletion. Dev Biol 146:278–291

    Article  CAS  Google Scholar 

  • Gavert N, Conacci-Sorrell M, Gast D, Schneider A, Altevogt P, Brabletz T, Ben-Ze’ev A (2005) L1, a novel target of beta-catenin signaling, transforms cells and is expressed at the invasive front of colon cancers. J Cell Biol 168:633–642

    Article  CAS  Google Scholar 

  • Harms BD, Bassi GM, Horwitz AR, Lauffenburger DA (2005) Directional persistence of EGF-induced cell migration is associated with stabilization of lamellipodial protrusions. Biophys J 88:1479–1488

    Article  CAS  Google Scholar 

  • Herren B, Garton KJ, Coats S, Bowen-Pope DF, Ross R, Raines EW (2001) ADAM15 overexpression in NIH3T3 cells enhances cell–cell interactions. Exp Cell Res 271:152–160

    Article  CAS  Google Scholar 

  • Hoang MV, Whelan MC, Senger DR (2004) Rho activity critically and selectively regulates endothelial cell organization during angiogenesis. Proc Natl Acad Sci USA 101:1874–1879

    Article  CAS  Google Scholar 

  • Huang C, Rajfur Z, Borchers C, Schaller MD, Jacobson K (2003) JNK phosphorylates paxillin and regulates cell migration. Nature 424:219–223

    Article  CAS  Google Scholar 

  • John GR, Chen L, Rivieccio MA, Melendez-Vasquez CV, Hartley A, Brosnan CF (2004) Interleukin-1beta induces a reactive astroglial phenotype via deactivation of the Rho GTPase-Rock axis. J Neurosci 24:2837–2845

    Article  CAS  Google Scholar 

  • Laurent-Matha V, Maruani-Herrmann S, Prebois C, Beaujouin M, Glondu M, Noel A, Alvarez-Gonzalez ML, Blacher S et al (2005) Catalytically inactive human cathepsin D triggers fibroblast invasive growth. J Cell Biol 168:489–499

    Article  CAS  Google Scholar 

  • Lee CC, Putnam AJ, Miranti CK, Gustafson M, Wang LM, Vande Woude GF, Gao CF (2004) Overexpression of sprouty 2 inhibits HGF/SF-mediated cell growth, invasion, migration, and cytokinesis. Oncogene 23:5193–5202

    Article  CAS  Google Scholar 

  • Lynch L, Vodyanik PI, Boettiger D, Guvakova MA (2005) Insulin-like growth factor I controls adhesion strength mediated by alpha5beta1 integrins in motile carcinoma cells. Mol Biol Cell 16:51–63

    Article  CAS  Google Scholar 

  • Maschler S, Wirl G, Spring H, Bredow DV, Sordat I, Beug H, Reichmann E (2005) Tumor cell invasiveness correlates with changes in integrin expression and localization. Oncogene 24:2032–2041

    Article  CAS  Google Scholar 

  • Miao H, Strebhardt K, Pasquale EB, Shen TL, Guan JL, Wang B (2005) Inhibition of integrin-mediated cell adhesion but not directional cell migration requires catalytic activity of EphB3 receptor tyrosine kinase. Role of Rho family small GTPases. J Biol Chem 280:923–932

    Article  CAS  Google Scholar 

  • Motegi S, Okazawa H, Ohnishi H, Sato R, Kaneko Y, Kobayashi H, Tomizawa K, Ito T et al (2003) Role of the CD47-SHPS-1 system in regulation of cell migration. EMBO J 22:2634–2644

    Article  CAS  Google Scholar 

  • Moyano JV, Maqueda A, Casanova B, Garcia-Pardo A (2003) Alpha4beta1 integrin/ligand interaction inhibits alpha5beta1-induced stress fibers and focal adhesions via down-regulation of RhoA and induces melanoma cell migration. Mol Biol Cell 14:3699–3715

    Article  CAS  Google Scholar 

  • Nishio T, Kawaguchi S, Yamamoto M, Iseda T, Kawasaki T, Hase T (2005) Tenascin-C regulates proliferation and migration of cultured astrocytes in a scratch wound assay. Neuroscience 132:87–102

    Article  CAS  Google Scholar 

  • Petridis AK, El-Maarouf A, Rutishauser U (2004) Polysialic acid regulates cell contact dependent neuronal differentiation of progenitor cells from the subventricular zone. Dev Dyn 230:675–684

    Article  CAS  Google Scholar 

  • Piccolo E, Innominato PF, Mariggio MA, Maffucci T, Iacobelli S, Falasca M (2002) The mechanism involved in the regulation of phospholipase Cgamma1 activity in cell migration. Oncogene 21:6520–6529

    Article  CAS  Google Scholar 

  • Pratt SJ, Epple H, Ward M, Feng Y, Braga VM, Longmore GD (2005) The LIM protein Ajuba influences p130Cas localization and Rac1 activity during cell migration. J Cell Biol 168:813–824

    Article  CAS  Google Scholar 

  • Raftopoulou M, Etienne-Manneville S, Self A, Nicholls S, Hall A (2004) Regulation of cell migration by the C2 domain of the tumor suppressor PTEN. Science 303:1179–1181

    Article  CAS  Google Scholar 

  • Ray RM, McCormack SA, Covington C, Viar MJ, Zheng Y, Johnson LR (2003) The requirement for polyamines for intestinal epithelial cell migration is mediated through Rac1. J Biol Chem 278:13039–13046

    Article  CAS  Google Scholar 

  • Robinet A, Fahem A, Cauchard JH, Huet E, Vincent L, Lorimier S, Antonicelli F, Soria C et al (2005) Elastin-derived peptides enhance angiogenesis by promoting endothelial cell migration and tubulogenesis through upregulation of MT1-MMP. J Cell Sci 118:343–356

    Article  CAS  Google Scholar 

  • Six EM, Ndiaye D, Sauer G, Laabi Y, Athman R, Cumano A, Brou C, Israel A, Logeat F (2004) The notch ligand Delta1 recruits Dlg1 at cell–cell contacts and regulates cell migration. J Biol Chem 279:55818–55826

    Article  CAS  Google Scholar 

  • Sun S, Wise J, Cho M (2004) Human fibroblast migration in three-dimensional collagen gel in response to noninvasive electrical stimulus. I. Characterization of induced three-dimensional cell movement. Tissue Eng 10:1548–1557

    CAS  Google Scholar 

  • Wadham C, Gamble JR, Vadas MA, Khew-Goodall Y (2003) The protein tyrosine phosphatase Pez is a major phosphatase of adherens junctions and dephosphorylates beta-catenin. Mol Biol Cell 14:2520–2529

    Article  CAS  Google Scholar 

  • Yarrow JC, Perlman ZE, Westwood NJ, Mitchison TJ (2004) A high-throughput cell migration assay using scratch wound healing, a comparison of image-based readout methods. BMC Biotechnol 4:21

    Article  Google Scholar 

  • Yoshida H, Cheng W, Hung J, Montell D, Geisbrecht E, Rosen D, Liu J, Naora H (2004) Lessons from border cell migration in the Drosophila ovary: a role for myosin VI in dissemination of human ovarian cancer. Proc Natl Acad Sci USA 101:8144–8149

    Article  CAS  Google Scholar 

  • Zhang L, Deng M, Parthasarathy R, Wang L, Mongan M, Molkentin JD, Zheng Y, Xia Y (2005) MEKK1 transduces activin signals in keratinocytes to induce actin stress fiber formation and migration. Mol Cell Biol 25:60–65

    Article  Google Scholar 

  • Zhu N, Lalla R, Eves P, Brown TH, King A, Kemp EH, Haycock JW, MacNeil S (2004) Melanoma cell migration is upregulated by tumour necrosis factor-alpha and suppressed by alpha-melanocyte-stimulating hormone. Brit J Cancer 90:1457–1463

    Article  CAS  Google Scholar 

  • Zhu X, Jiang J, Shen H, Wang H, Zong H, Li Z, Yang Y, Niu Z et al (2005) Elevated beta1,4-galactosyltransferase I in highly metastatic human lung cancer cells. Identification of E1AF as important transcription activator. J Biol Chem 280:12503–12516

    Article  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from NIH to D.S.G. (NS040317), to J.K. (NS049523), and to N.J.K. (HD042066). We gratefully acknowledge that the filter cube with custom excitation bandpass filters was provided by Chroma Technology Corporation. J. Fotos and V. Patel were awarded undergraduate research fellowships through grants to the Univ. of Delaware from the Howard Hughes Medical Institute and the Ronald E.␣McNair Program. We gratefully thank Dr. John Kappes for the GFP lentiviral vector construct and Dr. Marty Grumet for the NgCAM/L1 cDNA construct.

Author information

Author notes

  1. Norman J. Karin

    Present address: Pacific Northwest National Laboratory, 902 Battelle Blvd., Richland, WA, 99352, USA

Authors and Affiliations

  1. Department of Biological Sciences, University of Delaware, Wolf Hall, Newark, DE, 19716, USA

    Joseph S. Fotos, Vivek P. Patel, Norman J. Karin, Murali K. Temburni & Deni S. Galileo

  2. Chemistry and Biochemistry, University of Delaware, Newark, DE, 19716, USA

    Murali K. Temburni & John T. Koh

Authors
  1. Joseph S. Fotos
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Vivek P. Patel
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Norman J. Karin
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Murali K. Temburni
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. John T. Koh
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Deni S. Galileo
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Deni S. Galileo.

Additional information

Joseph S. Fotos and Vivek P. Patel contributed equally to this work

Rights and permissions

Reprints and permissions

About this article

Cite this article

Fotos, J.S., Patel, V.P., Karin, N.J. et al. Automated time-lapse microscopy and high-resolution tracking of cell migration. Cytotechnology 51, 7–19 (2006). https://doi.org/10.1007/s10616-006-9006-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10616-006-9006-7

Keywords

Search

Navigation