title: Binding affinities/avidities of antibody-antigen interactions: quantification and scale-up implications.
abstract: Bioaffinity interactions have been, and continue to be, successfully adapted from nature for use in separation and detection applications. It has been previously reported that the magnetophoretic mobility of labeled cells show a saturation type phenomenon as a function of the concentration of the free antibody-magnetic nanoparticle conjugate which is consistent with other reports of antibody-fluorophore binding. Starting with the standard antibody-antigen relationship, a model was developed which takes into consideration multi-valence interactions, and various attributes of flow cytometry (FCM) and cell tracking velocimetry (CTV) measurements to determine both the apparent dissociation constant and the antibody-binding capacity (ABC) of a cell. This model was then evaluated on peripheral blood lymphocytes (PBLs) labeled with anti CD3 antibodies conjugated to FITC, PE, or DM (magnetic nanoparticles). Reasonable agreements between the model and the experiments were obtained. In addition, estimates of the limitation of the number of magnetic nanoparticles that can bind to a cell as a result of steric hinderance was consistent with measured values of magnetophoretic mobility. Finally, a scale-up model was proposed and tested which predicts the amount of antibody conjugates needed to achieve a given level of saturation as the total number of cells reaches 10(10), the number of cells needed for certain clinical applications, such as T-cell depletions for mismatched bone marrow transplants.
authors.affiliation: Department of Chemical and Biomolecular Engineering, The Ohio State University, 140 W 19th Ave., Columbus, Ohio 43210, USA.
authors.forename: Huading
authors.initials: H
authors.lastname: Zhang
authors.forename: P Stephen
authors.initials: PS
authors.lastname: Williams
authors.forename: Maciej
authors.initials: M
authors.lastname: Zborowski
authors.forename: Jeffrey J
authors.initials: JJ
authors.lastname: Chalmers
mesh_terms: D000915:Antibody Affinity; D000936:Antigen-Antibody Complex / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D001710:Biotin / Q000031:analogs & derivatives* / Q000737:chemistry; D002469:Cell Separation / Q000379:methods*; D005434:Flow Cytometry; D007158:Immunologic Techniques; D008214:Lymphocytes; D008280:Magnetics; D008954:Models, Biological
chemical_list: D000936:Antigen-Antibody Complex; D001426:Bacterial Proteins; C066114:biotin-streptavidin complex; D001710:Biotin