A simple panning method for the selection of cell surface antigen transfectants

Mouse Ltk- (H-2k) cells that became reactive with anti-H-2d monoclonal antibodies (McAb) after transfection with DNA from H-2d-positive cells were isolated by an indirect panning method. The cells were transfected with DNA from a plasmid containing the selective thymidine kinase (tk) gene in addition to total cell DNA, so that a first selection could be carried out in hypoxanthine/aminopterin/thymidine (HAT) medium. The HAT-selected tk+ transfectants were incubated with anti-H-2d McAb, washed and transferred to dishes coated with purified anti-immunoglobulin (anti-Ig). This 2-step method of panning has the advantage that only the anti-Ig reagent requires purification. Transfected cells clearly reactive with either anti-Ld/Rd or anti-Kd McAb were isolated after only 1 or 2 cycles of panning.