Recombinant Fusion Proteins and Libraries from Immune Cell Repertoires

A method for preparing a plurality of recombinant T cell receptor expression constructs, comprising:

• providing a plurality of recombinant fusion polynucleotides each comprising a first polynucleotide encoding a first variable domain, a second polynucleotide encoding a second variable domain, and a linker polynucleotide linking the first and second polynucleotides;
• circularizing one or more of said plurality of recombinant fusion polynucleotides;
• inserting a third polynucleotide comprising a sequence encoding a promoter, a sequence encoding an internal ribosome entry site, or a sequence encoding a constant region between said first and second polynucleotide in each of said circularized recombinant fusion polynucleotides, thereby generating a plurality of recombinant T cell receptor expression constructs;
• expressing said plurality of recombinant T cell receptor expression constructs to display a plurality of recombinant fusion polypeptides on a plurality of cell surfaces;
• and
• enriching a subpopulation of said plurality of recombinant fusion polynucleotides encoding recombinant fusion polypeptides that can bind to an antigen by exposing said plurality of surfaces to said antigen and selecting based on binding between said antigen and one or more of said plurality of recombinant fusion polypeptides.