Biomedical subjects

John C Crocker

Publications and source records attributed to John C Crocker.

7 recordsLinked to original sources

PubMed2006

The role of F-actin and myosin in epithelial cell rheology.

Although actin and myosin are important contributors to cell-force generation, shape change, and motility, their contributions to cell stiffness and frequency-dependent rheology have not been conclusively determined. We apply several pharmacological interventions to cultured epithelial cells to elucidate the roles of actin and myosin in the mechanical response of cells and intracellular fluctuations. A suite of different methods is used to separately examine the mechanics of the deep cell interior and cortex, in response to depletion of intracellular ATP, depolymerization of F-actin, and inhibition of myosin II. Comparison of these results shows that F-actin plays a significant role in the mechanics of the cortical region of epithelial cells, but its disruption has no discernable effect on the rheology of the deeper interior. Moreover, we find that myosins do not contribute significantly to the rheology or ATP-dependent, non-Brownian motion in the cell interior. Finally, we investigate the broad distribution of apparent stiffness values reported by some microrheology methods, which are not observed with two-point microrheology. Based on our findings and a simple model, we conclude that heterogeneity of the tracer-cytoskeleton contacts, rather than the network itself, can explain the broad distribution of apparent stiffnesses.

Actins↗

PubMed2006

The consensus mechanics of cultured mammalian cells.

Although understanding cells' responses to mechanical stimuli is seen as increasingly important for understanding cell biology, how to best measure, interpret, and model cells' mechanical properties remains unclear. We determine the frequency-dependent shear modulus of cultured mammalian cells by using four different methods, both unique and well established. This approach clarifies the effects of cytoskeletal heterogeneity, ATP-dependent processes, and cell regional variations on the interpretation of such measurements. Our results clearly indicate two qualitatively similar, but distinct, mechanical responses, corresponding to the cortical and intracellular networks, each having an unusual, weak power-law form at low frequency. The two frequency-dependent responses we observe are remarkably similar to those reported for a variety of cultured mammalian cells measured with different techniques, suggesting it is a useful consensus description. Finally, we discuss possible physical explanations for the observed mechanical response.

Adenosine Triphosphate↗

PubMed2006

Engineering DNA-mediated colloidal crystallization.

DNA is a powerful and versatile tool for nanoscale self-assembly. Several researchers have assembled nanoparticles and colloids into a variety of structures using the sequence-specific binding properties of DNA. Until recently, however, all of the reported structures were disordered, even in systems where ordered colloidal crystals might be expected. We detail the experimental approach and surface preparation that we used to form the first DNA-mediated colloidal crystals, using 1 mum diameter polystyrene particles. Control experiments based on the depletion interaction clearly indicate that two standard methods for grafting biomolecules to colloidal particles (biotin/avidin and water-soluble carbodiimide) do not lead to ordered structures, even when blockers are employed that yield nominally stable, reversibly aggregating dispersions. In contrast, a swelling/deswelling-based method with poly(ethylene glycol) spacers resulted in particles that readily formed ordered crystals. The sequence specificity of the interaction is demonstrated by the crystal excluding particles bearing a noninteracting sequence. The temperature dependence of gelation and crystallization agree well with a simple thermodynamic model and a more detailed model of the effective colloidal pair interaction potential. We hypothesize that the surfaces yielded by the first two chemistries somehow hinder the particle-particle rolling required for annealing ordered structures, while at the same time not inducing a significant mean-force interaction that would alter the self-assembly phase diagram. Finally, we observe that particle crystallization kinetics become faster as the grafted-DNA density is increased, consistent with the particle-particle binding process being reaction, rather than diffusion limited.

Avidin↗

PubMed2005

Reversible self-assembly and directed assembly of DNA-linked micrometer-sized colloids.

We present a technique for the directed assembly and self-assembly of micrometer-scale structures based on the control of specific DNA linkages between colloidal particles. The use of DNA links combined with polymer brushes provides an effective way to regulate the range and magnitude of addressable forces between pairs (and further combinations) of different particles. We demonstrate that the autoassembly of alternate microbeads as well as their directed assembly, by using laser tweezers, is reversible. The key to reversibility is preventing the particles from falling into their van der Waals well at close distances. This goal is achieved by the use of adsorbed polymers that limit the number of DNA bridges to one to three between adjacent particles.

Base Sequence↗

PubMed2005

Swelling-based method for preparing stable, functionalized polymer colloids.

We describe a swelling-based method to prepare sterically stabilized polymer colloids with different functional groups or biomolecules attached to their surface. It should be applicable to a variety of polymeric colloids, including magnetic particles, fluorescent particles, polystyrene particles, PMMA particles, and so forth. The resulting particles are more stable in the presence of monovalent and divalent salt than existing functionalized colloids, even in the absence of any surfactant or protein blocker. While we use a PEG polymer brush here, the method should enable the use of a variety of polymer chemistries and molecular weights.

Colloids↗

PubMed2005

Colloidal interactions and self-assembly using DNA hybridization.

The specific binding of complementary DNA strands has been suggested as an ideal method for directing the controlled self-assembly of microscopic objects. We report the first direct measurements of such DNA-induced interactions between colloidal microspheres, as well as the first colloidal crystals assembled using them. The interactions measured with our optical tweezer method can be modeled in detail by well-known statistical physics and chemistry, boding well for their application to directed self-assembly. The microspheres' binding dynamics, however, have a surprising power-law scaling that can significantly slow annealing and crystallization.

Colloids↗

PubMed2002

Microrheology of polyethylene oxide using diffusing wave spectroscopy and single scattering.

Experiments investigating the local viscoelastic properties of a simple uncross-linked flexible polymer are performed on polyethylene oxide solutions in the semidilute regime using polystyrene beads of varying sizes and surface chemistry as probes. We measure the thermal motions of the beads to obtain the elastic and viscous moduli of our sample. Two different dynamic light scattering techniques, diffusing wave spectroscopy and quasielastic light scattering (QELS), are used to determine the dynamics of the probe particles. Diffusing wave spectroscopy probes the short time dynamics of the scatterers while QELS or single scattering measures the dynamics at larger times. This results in a larger frequency overlap of the data obtained from the microrheological techniques with the data obtained from the conventional bulk measurements. The moduli are estimated using a modified algebraic form of the generalized Stokes-Einstein equation. Comparison of microrheology with bulk measurements shows excellent similarity confirming the applicability of this method for simple, uncross-linked polymeric systems.

Journal Article↗