A novel method for localizing reporter fluorescent beads near the cell culture surface for traction force microscopy

Samantha G. Knoll, M. Yakut Ali, M. Taher A. Saif

Research output: Contribution to journalArticle

Abstract

PA gels have long been used as a platform to study cell traction forces due to ease of fabrication and the ability to tune their elastic properties. When the substrate is coated with an extracellular matrix protein, cells adhere to the gel and apply forces, causing the gel to deform. The deformation depends on the cell traction and the elastic properties of the gel. If the deformation field of the surface is known, surface traction can be calculated using elasticity theory. Gel deformation is commonly measured by embedding fluorescent marker beads uniformly into the gel. The probes displace as the gel deforms. The probes near the surface of the gel are tracked. The displacements reported by these probes are considered as surface displacements. Their depths from the surface are ignored. This assumption introduces error in traction force evaluations. For precise measurement of cell forces, it is critical for the location of the beads to be known. We have developed a technique that utilizes simple chemistry to confine fluorescent marker beads, 0.1 and 1 µm in diameter, in PA gels, within 1.6 μm of the surface. We coat a coverslip with poly-D-lysine (PDL) and fluorescent beads. PA gel solution is then sandwiched between the coverslip and an adherent surface. The fluorescent beads transfer to the gel solution during curing. After polymerization, the PA gel contains fluorescent beads on a plane close to the gel surface.

Original languageEnglish (US)
Article numbere51873
JournalJournal of Visualized Experiments
Issue number91
DOIs
StatePublished - Sep 16 2014

Keywords

  • Bioengineering
  • Cell culture surface
  • Cell mechanics
  • Fluorescent beads
  • Issue 91
  • Poly-D-lysine (PDL)
  • Polyacrylamide (PA) gel
  • Traction force microscopy

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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