Method for making variant secreted proteins with altered properties

A method for making a variant secretory protein retaining at least one desired binding property and eliminating at least one undesired binding property of the wild-type secretory protein comprising:

• (a) mutating the DNA encoding the selected wild-type protein thereby creating a library of related variant DNA molecules;
• (b) inserting each DNA molecule created in step (a) into an expression vector, wherein the vector comprises DNA encoding a transmembrane anchor domain thereby creating a library of vectors;
• (c) transfecting eukaryotic cells with the vectors of step (b);
• (d) culturing the cells of step (c) under conditions inducing the expression of the DNA to produce a chimeric fusion protein immobilized on the cell membrane;
• (e) contacting the cultured cells of step (d) with first and second reporter molecules capable of binding to different epitopes on the wild-type protein, each reporter molecule containing a fluorophore different from the fluorophore of the other reporter molecule, with said contacting taking place under conditions for which at least a portion of the cultured cells bind to the first or second reporter molecules;
• (f) sorting the contacted cells based on a desired binding pattern with the first or second reporter molecules;
• and
• (g) obtaining the variant proteins having the desired binding pattern from the sorted cells from (f).