Determination of the elastic moduli of thin samples and adherent cells using conical atomic force microscope tips

Abstract

The atomic force microscope can detect the mechanical fingerprints of normal and diseased cells at the single-cell level under physiological conditions1,2. However, atomic force microscopy studies of cell mechanics are limited by the ‘bottom effect’ artefact that arises from the stiff substrates used to culture cells. Because cells adhered to substrates are very thin3, this artefact makes cells appear stiffer than they really are4. Here, we show an analytical correction that accounts for this artefact when conical tips are used for atomic force microscope measurements of thin samples. Our bottom effect cone correction (BECC) corrects the Sneddon's model5, which is widely used to measure Young's modulus, E. Comparing the performance of BECC and Sneddon's model on thin polyacrylamide gels, we find that although Sneddon's model overestimates E, BECC yields E values that are thickness-independent and similar to those obtained on thick regions of the gel. The application of BECC to measurements on live adherent fibroblasts demonstrates a significant improvement on the estimation of their local mechanical properties.

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Figure 1: BECC removes the bottom-effect artefact in measurements performed on thin gel samples.
Figure 2: Indentations larger than 10% of the sample thickness result in a very large overestimaton of the elastic modulus.
Figure 3: BECC allows non-artefactual measurement of the local elastic moduli of adherent cells.
Figure 4: NIH3T3 cells display the greatest variability in E in regions that are <4 µm thick.

References

  1. 1

    Lee, G. Y. H. & Lim, C. T. Biomechanics approaches to studying human diseases. Trends Biotechnol. 25, 111–118 (2007).

    CAS  Article  Google Scholar 

  2. 2

    Costa, K. D. Single-cell elastography: probing for disease with the atomic force microscope. Dis. Markers 19, 139–154 (2003–2004).

    Article  Google Scholar 

  3. 3

    Lekka, M. & Laidler, P. Applicability of AFM in cancer detection. Nature Nanotech. 4, 72 (2009).

    CAS  Article  Google Scholar 

  4. 4

    Dimitriadis, E. K., Horkay, F., Maresca, J., Kachar, B. & Chadwick, R. Determination of elastic moduli of thin layers of soft material using the atomic force microscope. Biophys. J. 82, 2798–2810 (2002).

    CAS  Article  Google Scholar 

  5. 5

    Sneddon, I. N. Fourier Transforms Ch. 10 (McGraw-Hill, 1951).

    Google Scholar 

  6. 6

    Lekka, M. et al. Elasticity of normal and cancerous human bladder cells studied by scanning force microscopy. Eur. Biophys. J. 28, 312–316 (1999).

    CAS  Article  Google Scholar 

  7. 7

    Cross, S. E., Jin, Y-S., Rao, J. Y. & Gimzewski, J. K. Nanomechanical analysis of cells from cancer patients. Nature Nanotech. 2, 780–783 (2007).

    CAS  Article  Google Scholar 

  8. 8

    Cross, S. E. et al. AFM-based analysis of human metastatic cancer cells. Nanotechnology 19, 384003–384011 (2008).

    Article  Google Scholar 

  9. 9

    Li, Q. S., Lee, G. Y., Ong, C. N. & Lim, C. T. AFM indentation study of breast cancer cells. Biochem. Biophys. Res. Commun. 374, 609–613 (2008).

    CAS  Article  Google Scholar 

  10. 10

    Lekka, M. et al. Cancer cell recognition—mechanical phenotype. Micron 44, 1259–1266 (2012).

    Article  Google Scholar 

  11. 11

    Prabhune, M. et al. Comparison of mechanical properties of normal and malignant thyroid cells. Micron 43, 1267–1272 (2012).

    Article  Google Scholar 

  12. 12

    Kuznetsova, T. G. et al. Atomic force microscopy probing of cell elasticity. Micron 38, 824–833 (2007).

    CAS  Article  Google Scholar 

  13. 13

    Cross, S. E., Jin, Y-S., Rao, J. & Gimzewski, J. K. Applicability of AFM in cancer detection. Nature Nanotech. 4, 72–73 (2009).

    CAS  Article  Google Scholar 

  14. 14

    Rico, F. et al. Probing mechanical properties of living cells by atomic force microscopy with blunted pyramidal cantilever tips. Phys. Rev. E 72, 021914 (2005).

    Article  Google Scholar 

  15. 15

    Kang, I. et al. Changes in the hyperelastic properties of endothelial cells induced by tumor necrosis factor-α. Biophys. J. 94, 3273–3285 (2008).

    CAS  Article  Google Scholar 

  16. 16

    Kihara, T., Haghparast, S. M. A., Shimizu, Y., Yuba, S. & Miyake, J. Physical properties of mesenchymal stem cells are coordinated by the perinuclear actin cap. Biochem. Biophys. Res. Commun. 409, 1–6 (2011).

    CAS  Article  Google Scholar 

  17. 17

    McElfresh, M. et al. Combining constitutive materials modeling with atomic force microscopy to understand the mechanical properties of living cells. Proc. Natl Acad. Sci. USA 99 (Suppl 2), 6493–6497 (2002).

    CAS  Article  Google Scholar 

  18. 18

    Alcaraz, J. et al. Microrheology of human lung epithelial cells measured by atomic force microscopy. Biophys. J. 84, 2071–2079 (2003).

    CAS  Article  Google Scholar 

  19. 19

    Raman, A. et al. Mapping nanomechanical properties of live cells using multi-harmonic atomic force microscopy. Nature Nanotech. 6, 809–814 (2011).

    CAS  Article  Google Scholar 

  20. 20

    Pogoda, K. et al. Depth-sensing analysis of cytoskeleton organization based on AFM data. Eur. Biophys. J. 41, 79–87 (2012).

    CAS  Article  Google Scholar 

  21. 21

    Petrie, R. J., Gavara, N., Chadwick, R. S. & Yamada, K. M. Nonpolarized signaling reveals two distinct modes of 3D cell migration. J. Cell Biol. 197, 439–455 (2012).

    CAS  Article  Google Scholar 

  22. 22

    Gavara, N. & Chadwick, R. S. Noncontact microrheology at acoustic frequencies using frequency-modulated atomic force microscopy. Nature Methods 7, 650–654 (2010).

    CAS  Article  Google Scholar 

  23. 23

    Wong, J. Y., Velasco, A., Rajagopalan, P. & Pham, Q. Directed movement of vascular smooth muscle cells on gradient-compliant hydrogels. Langmuir 19, 1908–1913 (2003).

    CAS  Article  Google Scholar 

  24. 24

    Butt, H. J. & Jaschke, M. Calculation of thermal noise in atomic force microscopy. Nanotechnology 6, 1–7 (1995).

    Article  Google Scholar 

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Acknowledgements

The authors thank R. Sunyer, V. Luo and K.M. Yamada for critical input. This work was supported by the Intramural Program of the US National Institute of Deafness and Other Communication Disorders.

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N.G. conceived, designed and performed the experiments and analysed the data. R.S.C. developed the BECC correction. N.G. and R.S.C. co-wrote the paper.

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Correspondence to Richard S. Chadwick.

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The authors declare no competing financial interests.

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Gavara, N., Chadwick, R. Determination of the elastic moduli of thin samples and adherent cells using conical atomic force microscope tips. Nature Nanotech 7, 733–736 (2012). https://doi.org/10.1038/nnano.2012.163

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