John A. Quinn

Restricted Transport in Small Pores: A Model for Steric Exclusion and Hindered Particle Motion

The basic hydrodynamic equations governing transport in submicron pores are reexamined. Conditions necessary for a simplified, one-dimensional treatment of the diffusion/convection process are established. Steric restrictions and Brownian motion are incorporated directly into the resulting model. Currently available fluid mechanical results are used to evaluate an upper limit on hindered diffusion; this limit is valid for small particle-to-pore ratios. Extensions of the analysis are shown to depend on numerical solutions of the related hydrodynamic problem, that of asymmetrical particle motion in a bounded fluid.

Receptor-mediated adhesion phenomena. Model studies with the Radical-Flow Detachment Assay

Receptor-mediated cell adhesion phenomena play a vital role in many physiological and biotechnology-related processes. To investigate the physical and chemical factors that influence the cell/surface interaction, we have used a radial flow device, a so-called Radial-Flow Detachment Assay (RFDA). The RFDA allows us to make direct observations of the detachment process under specified experimental conditions. In results reported here, we have studied the detachment of receptor-coated latex beads (prototype cells) from ligand-coated glass surfaces. The receptors and ligands used in this work are complementary antibodies. The beads enable us to examine several aspects of the adhesion process with particles having uniform properties that can be varied systematically. Advantages of the RFDA are many, especially direct observation of cell detachment over a range of shear stresses with quantitative measurement of the adhesive force. We focus our studies on the effects of ligand and receptor densities, along with the influence of pH and ionic strength of the medium. These data are analyzed with a mathematical model based on the theoretical framework of Bell, G. I. (1978. Science [Wash. DC]. 200:618-627) and Hammer, D. A. and D. A. Lauffenburger (1987. Biophys. J. 52:475-487). We demonstrate experimental validation of a theoretical expression for the critical shear stress for particle detachment, and show that it is consistent with reasonable estimates for the receptor-ligand bond affinity.

Receptor-mediated cell attachment and detachment kinetics. I. Probabilistic model and analysis

The kinetics of receptor-mediated cell adhesion to a ligand-coated surface play a key role in many physiological and biotechnology-related processes. We present a probabilistic model of receptor-ligand bond formation between a cell and surface to describe the probability of adhesion in a fluid shear field. Our model extends the deterministic model of Hammer and Lauffenburger (Hammer, D.A., and D.A. Lauffenburger. 1987. Biophys. J. 52:475-487) to a probabilistic framework, in which we calculate the probability that a certain number of bonds between a cell and surface exists at any given time. The probabilistic framework is used to account for deviations from ideal, deterministic behavior, inherent in chemical reactions involving relatively small numbers of reacting molecules. Two situations are investigated: first, cell attachment in the absence of fluid stress; and, second, cell detachment in the presence of fluid stress. In the attachment case, we examine the expected variance in bond formation as a function of attachment time; this also provides an initial condition for the detachment case. Focusing then on detachment, we predict transient behavior as a function of key system parameters, such as the distractive fluid force, the receptor-ligand bond affinity and rate constants, and the receptor and ligand densities. We compare the predictions of the probabilistic model with those of a deterministic model, and show how a deterministic approach can yield some inaccurate results; e.g., it cannot account for temporally continuous cell attach mentor detachment, it can underestimate the time needed for cell attachment, it can overestimate the time required for cell detachment for a given level of force, and it can overestimate the force necessary for cell detachment.

Receptor-mediated cell attachment and detachment kinetics. II. Experimental model studies with the radial-flow detachment assay.

Quantitative information regarding the kinetics of receptor-mediated cell adhesion to a ligand-coated surface are crucial for understanding the role of certain key parameters in many physiological and biotechnology-related processes. Here, we use the probabilistic attachment and detachment models developed in the preceding paper to interpret transient data from well-defined experiments. These data are obtained with a simple model cell system that consists of receptor-coated latex beads (prototype cells) and a Radial-Flow Detachment Assay (RFDA) using a ligand-coated glass disc. The receptors and ligands used in this work are complementary antibodies. The beads enable us to examine transient behavior with particles that possess fairly uniform properties that can be varied systematically, and the RFDA is designed for direct observation of adhesion to the ligand-coated glass surface over a range of shear stresses. Our experiments focus on the effects of surface shear stress, receptor density, and ligand density. These data provide a crucial test of the probabilistic framework. We show that these data can be explained with the probabilistic analyses, whereas they cannot be readily interpreted on the basis of a deterministic analysis. In addition, we examine transient data on cell adhesion reported from other assays, demonstrating the consistency of these data with the predictions of the probabilistic models.

Carbon dioxide transport through enzymatically active synthetic membranes

The facilitated transport of CO2 through thin liquid membranes, Millipore filter membranes, and cross-linked protein membranes has been investigated using a tracer 14CO2 technique. Both the uncatalyzed reactions and the enzymatic reactions catalyzed by carbonic anhydrase in homogeneous solution and immobilized in several membrane configurations were studied. The steady-state transport data were reduced in terms of an analytical model for simultaneous reaction and diffusion which permitted the direct determination of the diffusional and enzyme kinetic parameters. The experimental method illustrates a powerful technique for measuring rapid reaction kinetics. In addition, a general multi-layer membrane model is developed which is capable of treating membrane kinetic heterogeneities. The multi-layer model yields a useful definition for the reaction boundary layer and provides a comparison among different membrane kinetic configurations for membrane design purposes.

Diffusion-induced banding of colloid particles via diffusiophoresis

Etude de la diffusiophorese de particules non electrolytiques. Application a la diffusiophorese de particules de Latex dans le percoll ou des solutions de dextran

Diffusion-induced banding of colloid particles via diffusiophoresis: 2. Non-electrolytes

Abstract We have extended the stopped-flow diffusion cell (SFDC) technique (Staffeld, P. O., and Quinn, J. A., J. Colloid Interface Sci. 130, 69 (1989) to measurements of nonelectrolyte diffusiophoresis using 1-μm diameter latex particles in gradients of Dextran and Percoll. Dextran, a slightly branched nonionic polymer, and Percoll, a negatively charged polymer coated silica particle, interact with latex via steric exclusion at the latex surface. Anderson and Prieves theory of nonelectrolyte diffusiophoresis is used to interpret the experimental measurements. Interaction parameters determined from our measurements are physically realistic and consistent with theoretical predictions.

Membrane Reactors in Bioprocessing

Engineering research on membranes has had two prime objectives, the first and by far the more extensively investigated being the development of membranes for separation processes.’” In parallel with this work, however, other investigators have considered microporous membranes as solid-phase supports to which enzymes oi’whole cells might be attached for the purpose of catalyzing a bioconversion.M These types of enzymatically active membranes, which we refer to as “reactive membranes,” have been operated in both diffusive and convective modes. Since membranes can effect both biocatalysis and separation, and since these two operations are inextricably related in any biochemical process, we were intrigued by the possibility of combining these functions in a single membrane device, the so-called membrane bioreactor, in hopes of achieving novel and powerful device performance.

Adsorption and elution of extracellular matrix proteins on non-tissue culture polystyrene petri dishes

Abstract We have devised a simple procedure for directly measuring the adsorption of fibrinogen, type IV collagen, and fibronectin on non-tissue culture polystyrene petri dishes using proteins labeled with 125I and removable dish sections. Our approach provides a rapid method for obtaining quantitative adsorption isotherms for these large extracellular matrix (ECM) proteins in situations applicable for studies of cell culture and cell adhesion migration . We found that the surface density of adsorbed ECM protein depends not only on solution protein concentration but also on ECM protein type, with roughly one and two orders of magnitude more fibronectin and type IV collagen, respectively, adsorbing at an equivalent solution concentration compared to fibrinogen. Adsorption isotherms for each of these proteins were compared with theoretical bounds for monolayer density based on random sequential adsorption and molecular close-packing. We also observed that exposure to 1% sodium dodecyl sulfate in 3 M NaOH for 25 h was effective at eluting fibrinogen from our dishes over a wide range of protein concentration, but that this same detergent treatment was ineffective at completely desorbing type IV collagen adsorbed at high density. Our results, useful to researchers examining the role of substratum-bound molecules in the control of cell behavior, demonstrate that estimates of the adsorbed molecular density of ECM proteins obtained from indirect methods, such as elution and enzyme-linked immunosorbent assays, or inferred from solution coating concentrations may provide erroneous estimates of the true number of molecules actually adsorbed.

Phase-transfer catalysis in a membrane reactor

Abstract Many contacting problems associated with conventional phase-transfer catalysis operations can be eliminated by using a membrane reactor. Instead of the mixer/settler sequence of the conventional reactor, the aqueous and organic streams are contacted across a porous, hydrophobic membrane—the role of the membrane is to stabilize the oil/water interface at a fixed position while affording efficient transfer of the phase-transfer agent. General mathematical models are presented for the behaviour of the membrane reactor and important dimensionless groups are identified. Model predictions are verified with an experimental demonstration using a flat sheet, thin film reactor. In addition to superior reactor performance, the fact that the membrane device can be operated under closely controlled and completely identifiable mass-transfer characteristics provides a convenient laboratory tool for investigating the kinetics of phase-transfer reactions, an increasingly important class of industrial synthesis reactions.