Isolation of cell surface-specific human monoclonal antibodies using phage display and magnetically-activated cell sorting: applications in immunohematology

@article{citeulike:2615667, abstract = {A method is described for the isolation of filamentous phage-displayed human monoclonal antibodies directed at unpurifiable cell surface-expressed molecules. To optimize the capture of antigen-specific phage and minimize the binding of irrelevant phage antibodies, a simultaneous positive and negative selection strategy is employed. Cells bearing the antigen of interest are pre-coated with magnetic beads and diluted into an excess of unmodified antigen-negative cells. Following incubation of the cell admixture with a Fab/phage library, the antigen-positive cell population is retrieved using magnetically-activated cell sorting and antigen-specific Fab/phage are eluted and propagated in bacterial culture. Utilizing this protocol with magnetically-labeled Rh(D)-positive and excess unlabeled Rh(D)-negative human red blood cells and a Fab/phage library constructed from human peripheral blood lymphocytes, dozens of unique clinically-useful [gamma]1[kappa] and [gamma]1[lambda] anti-Rh(D) antibodies were isolated from a single alloimmunized individual. This cell-surface selection method is readily adaptable for use in other systems, such as for the identification of putative tumor-specific antigens and provides a rapid (<1 month), high-yield approach for isolating self-replicative antibody reagents directed at novel or conformationally-dependent cell-surface epitopes.}, author = {Siegel, Don L. and Chang, Tylis Y. and Russell, Shari L. and Bunya, Vatinee Y. }, citeulike-article-id = {2615667}, doi = {10.1016/S0022-1759(97)00087-2}, journal = {Journal of Immunological Methods}, keywords = {hematology, methods}, month = {August}, number = {1-2}, pages = {73--85}, posted-at = {2008-03-31 11:05:13}, priority = {2}, title = {Isolation of cell surface-specific human monoclonal antibodies using phage display and magnetically-activated cell sorting: applications in immunohematology}, url = {http://dx.doi.org/10.1016/S0022-1759(97)00087-2}, volume = {206}, year = {1997} }

TY - JOUR ID - citeulike:2615667 L3 - citeulike-article-id:2615667 TI - Isolation of cell surface-specific human monoclonal antibodies using phage display and magnetically-activated cell sorting: applications in immunohematology JF - Journal of Immunological Methods VL - 206 IS - 1-2 SP - 73 EP - 85 N2 - A method is described for the isolation of filamentous phage-displayed human monoclonal antibodies directed at unpurifiable cell surface-expressed molecules. To optimize the capture of antigen-specific phage and minimize the binding of irrelevant phage antibodies, a simultaneous positive and negative selection strategy is employed. Cells bearing the antigen of interest are pre-coated with magnetic beads and diluted into an excess of unmodified antigen-negative cells. Following incubation of the cell admixture with a Fab/phage library, the antigen-positive cell population is retrieved using magnetically-activated cell sorting and antigen-specific Fab/phage are eluted and propagated in bacterial culture. Utilizing this protocol with magnetically-labeled Rh(D)-positive and excess unlabeled Rh(D)-negative human red blood cells and a Fab/phage library constructed from human peripheral blood lymphocytes, dozens of unique clinically-useful [gamma]1[kappa] and [gamma]1[lambda] anti-Rh(D) antibodies were isolated from a single alloimmunized individual. This cell-surface selection method is readily adaptable for use in other systems, such as for the identification of putative tumor-specific antigens and provides a rapid (<1 month), high-yield approach for isolating self-replicative antibody reagents directed at novel or conformationally-dependent cell-surface epitopes. KW - hematology KW - methods AU - Siegel, Don AU - Chang, Tylis AU - Russell, Shari AU - Bunya, Vatinee PY - 1997/08/07/ UR - http://dx.doi.org/10.1016/S0022-1759(97)00087-2 ER -