21^st New England Workshop on Complex Fluids
December 3, 2004
Materials Research Science and Engineering Center
Harvard University
  Agenda

Invited Talks
 

9:30 – 10:00 a.m.
Anubhav Tripathi, Brown University
Microfluidic Separation of Proteins in Semi-dilute Polymer Solutions
 
We present a systematic study of the electrophoretic migration of protein fragments (10- 200 kDa) in dilute-polymer solutions using microfluidic chips. The electrophoretic mobility and dispersion of protein samples were measured in a series of monodisperse polydimethylacrylamide (PDMA) polymers of different molecular weights (243, 443 & 764 kDa, polydispersivity index = 2) with varying concentrations. Experiments were also performed in PDMA solutions with a polydispersivity index of about 5 and in mixtures of polymers with different molecular weights. The polymers solutions were characterized using rheometry and optical techniques. Prior to its loading onto the microchip, the polymer solution was mixed with known concentrations of SDS surfactant and staining dye.  The SDS-denatured protein samples were electrokinetically injected, separated and detected in the microchip using electric fields ranging from 100 to 300 V/cm.

Our results show that the electrophoretic mobility of protein fragments decreases exponentially with the concentration cof the polymer solution. The mobility was found to decrease logarithmically with the molecular weight of the protein fragment. In addition, the mobility was found to be independent of the electric field in the separation channel. The results appear to suggest that the denatured protein molecules migrate as rigid rod-like molecules. The migration mechanism for protein molecules will be discussed using polymer physics and hydrodynamic drag arguments.
The protein migration was found to depend strongly on the SDS concentration in the polymer solution. Moreover, the optimal concentration of SDS depends on the polydispersity of the PDMA polymer. Specifically, repeated injection of protein samples through a single load of polymer solution was achieved using the optimized value of the SDS concentration. A simple mass balance analysis is used to explain the behavior. The results from experiments using a matrix composed of polyethylene oxide (PEO) and PDMA-PEO blends of varying composition will also be presented.  
 
10:00 – 10:30 a.m.
Peter Schall, Harvard University
Growth and Deformation of Colloidal Crystals and Glasses

Plastic deformation in atomic crystals is governed by dislocations — line defects in the crystalline lattice. Understanding how these dislocations propagate, multiply, and interact is central for understanding the plastic response of a crystalline material to an applied stress. Unlike for crystals, not much is known about the deformation mechanism of glasses, and the microscopic processes leading to macroscopic deformation are still highly speculative.

We use colloidal crystals and glasses as models to study the behavior of their atomic counterparts under applied stress. We use confocal microscopy to determine the position of the individual particles and to study defect propagation on the particle scale. The colloidal crystals exhibit dislocations that show remarkable similarities to dislocations in atomic crystals. The slow time scale of the colloidal suspension allows us to directly observe the nucleation of dislocations as the crystal is deformed. We have built a laser diffraction microscope, which is inspired by a transmission electron microscope used to study dislocations in hard materials, to image the strain field of the dislocation defects. This technique enables us to study dislocation motion and dislocation interaction on a much larger length scale.

In the amorphous suspension, we are able to follow the motion of the individual particles and identify local shear events that give rise to the macroscopic deformation. Recent results indicate a correlation between the location of shear events and regions of low local particle density or high free volume.

1:30–2:00
Peter G. Vekilov, University of Houston
Phase Transitions in Protein Solutions
 
Dense liquid, gel-like, and solid, ordered in three, two, or one dimension, or completely disordered phases form in protein solutions and underlie physiological and patho -physiological, laboratory, and technological processes.

Two aspects of the phase transitions will be discussed.

The first one is the role of water, structured at the hydrophobic and hydrophilic patches on the surface of the protein molecules.  Examples will be provided illustrating that this structuring often determines the entropy and enthalpy balance of the phase transition, leads to unusual intermolecular interaction potentials with one or more outlying maxima, which severely affect the phase diagrams, and that the dynamics of destruction of the water shell is the major determinant of the kinetics of association of molecules into solid phases.  Because of the water structuring, the fastest pathway of nucleation of ordered solid phases is not the one with the lowest free-energy barriers.

The second aspect is the interaction between the phases.  Examples from the nucleation of two types of ordered solid phases: three-dimensional crystals and the polymers of sickle cell hemoglobin, which have one-dimensional translational symmetry, show that nucleation proceeds via a disordered liquid-like intermediate. In crystal nucleation, the structuring of the intermediate is the rate determining step in the nucleation process, while in the nucleation of the HbS polymers, the formation of the intermediate determines the overall nucleation rate.
 

 
2:00 – 2:30 p.m.
Krystyn J. Van Vliet, Massachusetts Institute of Technology
Guiding and Determining Cell State through Chemomechanical Contact
 
Living cells can be idealized as chemomechanically coupled material systems, in that cells respond actively to changes in mechanical state through biomolecular synthesis, and likewise respond actively to changes in biochemical state through mechanical actuation. This feature of cells not only enables all biological processes, but also presents opportunities to guide cell development and to engineer robust transducers based on this coupling. Here, we discuss experimental means to manipulate and characterize the cell as an "active material" via nanomechanics. First, we will demonstrate the effects of cyclic mechanical strain on vascular endothelial cell phenotype. Second, we will discuss robust polyelectrolyte substrata by which we can independently control the mechanical compliance and biomolecular surface profile, such that the mechanical and chemical environments to which can be decoupled. Using this system, we show that phenotypic differentiation of vascular endothelial cells can be manipulated directly via the synthesis-dependent compliance of polymeric substrata. Finally, we present an approach to track the effects of external stimuli on the living cell, through chemomechanical mapping of molecules at or near the cell surface.
 
4:00 p.m.
Clare M. Waterman- Storer, The Scripps Research Institute, La Jolla ,CA
Cytoskeletal Systems Integration in Cell Migration
 
The directed locomotion of vertebrate tissue cells involves spatiotemporal coordination of protrusion, adhesion, and contraction.  This necessitates complex and dynamic interactions between cytoskeletal systems and the extracellular environment.  My lab uses quantitative microscopy of protein dynamics in living cells and in vitro biochemistry to understand how seemingly distinct cytomechanical systems are integrated with one another to promote the polarized morphogenic activity that drives directed cell movement.  My lab aims to answer questions such as how the microtubule and actin cytoskeletons interact to polarize a motile cell or how the acto -myosin contractile system interfaces with the extracellular matrix via focal adhesions to generate traction force that drives cell movement.  To aid our studies of molecular dynamics, we pioneered a method called quantitative Fluorescent Speckle Microscopy ( qFSM ), which allows quantitative analysis of the dynamics of and interactions between proteins within macromolecular assemblies such as the cytoskeleton and focal adhesions in living cells.  
 

Updated 12.2.04