Magnetic nanoparticle–mediated massively parallel mechanical modulation of single-cell behavior

ABSTRACT We report a technique for generating controllable, time-varying and localizable forces on arrays of cells in a massively parallel fashion. To achieve this, we grow magnetic

nanoparticle–dosed cells in defined patterns on micromagnetic substrates. By manipulating and coalescing nanoparticles within cells, we apply localized nanoparticle-mediated forces

approaching cellular yield tensions on the cortex of HeLa cells. We observed highly coordinated responses in cellular behavior, including the p21-activated kinase–dependent generation of

active, leading edge–type filopodia and biasing of the metaphase plate during mitosis. The large sample size and rapid sample generation inherent to this approach allow the analysis of cells

at an unprecedented rate: in a single experiment, potentially tens of thousands of cells can be stimulated for high statistical accuracy in measurements. This technique shows promise as a

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kinase. _Chem. Biol._ 15, 322–331 (2008). Article  CAS  Google Scholar  Download references ACKNOWLEDGEMENTS This work was partially supported through the US National Institutes of Health

Director's New Innovator Award (1DP2OD007113). The authors thank M. Bachman and N. Gunn (University of California, Irvine) for samples of PSR; J. Harrison, M. Glickman and I. Goldberg

for assistance with the permalloy electroplating bath; members of the UCLA Advanced Light Microscopy Spectroscopy facility for assistance with confocal microscopy; K. Lin for high-speed

imaging assistance; I. Williams for running FACS; and engineers of the UCLA Nanolab for processing assistance. AUTHOR INFORMATION AUTHORS AND AFFILIATIONS * Department of Bioengineering,

University of California, Los Angeles (UCLA), Los Angeles, California, USA Peter Tseng, Jack W Judy & Dino Di Carlo * Department of Electrical Engineering, UCLA, Los Angeles, California,

USA Peter Tseng & Jack W Judy * California NanoSystems Institute, UCLA, Los Angeles, California, USA Jack W Judy & Dino Di Carlo * Jonsson Comprehensive Cancer Center, UCLA, Los

Angeles, California, USA Dino Di Carlo Authors * Peter Tseng View author publications You can also search for this author inPubMed Google Scholar * Jack W Judy View author publications You

can also search for this author inPubMed Google Scholar * Dino Di Carlo View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS P.T. and D.D.C.

contributed to the initial concept. P.T. and J.W.J. contributed to the fabrication design. D.D.C. and P.T. designed the integration of magnetic elements and single cells. P.T. developed the

final fabrication and cell-patterning protocols. P.T. fabricated the micromagnetic slides and conducted the cell experiments. J.W.J. and P.T. discussed the finite-element simulation. P.T.

designed the numerical analysis flow. P.T. and D.D.C. discussed and analyzed the numerical results. All authors wrote the manuscript. CORRESPONDING AUTHOR Correspondence to Dino Di Carlo.

ETHICS DECLARATIONS COMPETING INTERESTS The authors declare no competing financial interests. SUPPLEMENTARY INFORMATION SUPPLEMENTARY TEXT AND FIGURES Supplementary Figures 1–9 (PDF 1355 kb)

MAGNETIC FLUORESCENT NANOPARTICLES COALESCE QUICKLY UNDER HIGH MAGNETIC FIELD STIMULATION (RESIN THICKNESS IS 0.5 ΜM). Nanoparticle assembly occurs over a period of ~30 min at the left edge

of the cell. As nanoparticles begin coalescing at the membrane edge, small clusters of nanoparticles enter temporary filopodial protrusions that extend beyond the edge. This effect

continues until around the 1-h mark, when the cell membrane begins to yield under the high tension until finally the nanoparticle cluster exhibits a 'pull-in' instability, and the

entire nanoparticle cluster protrudes from the edge of the cell membrane. Cell cytoplasm is labeled with calcein AM. (MOV 1472 kb) HIGH-SPEED IMAGES OF MAGNETIC BEADS MANIPULATED BY

INDIVIDUAL FERROMAGNETIC MICROMAGNETS. Video displays the trajectories of two magnetic beads moving along the substrate surface towards the magnetized elements using resins with thicknesses

of 2.5 and 5.3 μm, respectively. (AVI 970 kb) CONFOCAL MICROSCOPY _Z_ SLICES OF A SINGLE CELL UNDER MODERATE MAGNETIC NANOPARTICLE–MEDIATED TENSION. At the _z_ planes where the cell membrane

is under magnetic nanoparticle–induced tension, local effects are observed including (i) local deformation of the flanking stress fiber caused by the applied mechanical tension and (ii)

flanking actin-rich protrusions emanating from the regions of highest mechanical deformation. Positive myosin-X staining at the tips of protrusions indicate induced active, ECM-attachable

filopodia. (AVI 479 kb) CONFOCAL MICROSCOPY _Z_ SLICES SHOWING THE RICH BAND OF STRESS FIBER–LOCALIZED PHOSPHO-PAK PROGRESSING THROUGH THE CORTICAL REGIONS OF HIGH DEFORMATION. This

colocalization occurs whether or not the particular cell is expressing a high filopodial asymmetry and is distinct at regions directly above where the nanoparticles are localized. (AVI 1042

kb) CELL DIVIDING ALONG THE AXIS OF FORCE APPLICATION. The time-lapse video shows a single cell adhering to an I-shaped fibronectin pattern as it divides under high nanoparticle-induced

tension. As the cell undergoes and completes mitosis, the cell divides biased in the direction of the force generated by the magnetic nanoparticles. Upon successful division, both cells

adhere and move normally. The nanoparticles remain only in one of the daughter cells. (MOV 936 kb) RIGHTS AND PERMISSIONS Reprints and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Tseng,

P., Judy, J. & Di Carlo, D. Magnetic nanoparticle–mediated massively parallel mechanical modulation of single-cell behavior. _Nat Methods_ 9, 1113–1119 (2012).

https://doi.org/10.1038/nmeth.2210 Download citation * Received: 22 April 2012 * Accepted: 06 September 2012 * Published: 14 October 2012 * Issue Date: November 2012 * DOI:

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