title: Mass transfer in the biomolecular binding of a target against probe molecules on the surface of microbeads sequestered in wells in a microfluidic cell.
abstract: Diagnostic tools which screen the binding interactions of a protein target against a display of biomolecular probes to identify molecules which bind the target are central to cell proteomic studies, and to diagnostic assays. Here, we study a microfluidic design for screening interactions in which the probe molecules are hosted on microbeads sequestered in wells arranged at the bottom of a microfluidic flow channel. Assays are undertaken by streaming an analyte solution with a fluorescently labelled target through the cell, and identifying the fluorescing beads. Numerical simulations are first constructed for the analyte flow over the microbeads in the well array, and the increase in the target concentration on the microbead surface. The binding profile is expressed as a function of the ratio of the convective to the diffusive transport rates (Peclet number or Pe), and the ratio of the kinetic to the diffusive rates (Damkohler number, Da). For any Pe, as Da becomes small enough, the transport is determined by the intrinsic kinetic binding rate. As Pe increases, a thin concentration boundary layer develops over the top surface of the microbead because of the convective flow, and target binds more rapidly. However, the relatively stagnant layers of liquid in the well provide a diffusion barrier which slows the target transport, and for any Da and Pe the transport is slower than equivalent patches of probes arranged on the channel wall. Experiments are also undertaken at high Pe, using the binding of fluorescently labelled NeutrAvidin as a target to probes of its binding partner, biotin, on the microbead surface. The binding profile is compared to the simulations to measure the kinetic rate constant, and this comparison shows that the transport in the cell is not kinetically limited because of the diffusion barriers created by the stagnant liquid layer in the well. Simulations and experiments on microbeads which are only partially recessed in the well demonstrate an increase in the mass transfer rate as more of the microbead surface intersects the flow and the diffusion limitation due to the stagnant layer of liquid surrounding the bottom part of the microbead is minimized.
authors.affiliation: Levich Institute and Department of Chemical Engineering, The City College of the City University of New York, New York, New York 10031, USA. charles@chemail.engr.ccny.cuny.edu.
authors.forename: Thomas F
authors.initials: TF
authors.lastname: Leary
authors.forename: Setareh
authors.initials: S
authors.lastname: Manafirasi
authors.forename: Charles
authors.initials: C
authors.lastname: Maldarelli
mesh_terms: D001360:Avidin / Q000737:chemistry / Q000378:metabolism; D001710:Biotin / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry*; D007700:Kinetics; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D008856:Microscopy, Fluorescence; D008863:Microspheres; D008962:Models, Theoretical; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D013499:Surface Properties
chemical_list: D005456:Fluorescent Dyes; D011506:Proteins; D001360:Avidin; D001710:Biotin