Ligand screening and discovery

摘要

Disclosed is a method that includes: (i) providing a plurality of initial nucleic acid cassettes that include: a) a first coding region encoding a first immunoglobulin variable domain, b) a second coding region encoding a second immunoglobulin variable domain, and c) a ribosomal binding site disposed between the first and second coding regions for translation of the second polypeptide in a first expression system, wherein the first and second coding regions are in the same translational orientation; (ii) modifying each nucleic acid cassette of the plurality in a single reaction mixture so that it is functional in a second expression system, wherein the first and second region remain physically attached during the modifying; (iii) introducing each modified nucleic acid cassette into a mammalian cell to produce a mixture of transfected cells; and (iv) expressing each modified nucleic acid cassette in the transfected cells.

主权项

1. A method for evaluating display library members, the method comprising:
providing a first plurality of nucleic acid library members, each library member encoding a first immunoglobulin or function fragment thereof that is displayed on the surface of a prokaryotic cell, wherein each cassette comprises a first nucleic acid sequence encoding a first immunoglobulin variable domain, a second nucleic acid sequence encoding a second immunoglobulin variable domain, and a ribosomal binding site disposed between the first and second nucleic acid sequences, wherein the first and second nucleic acid sequences are in the same translational orientation and can be transcribed as a single transcript; determining an assessment for each library member with respect to a property; sorting information about the assessments of the library members in a computer database; filtering the information to identify a subset of the library members; and reformatting each library member in a single reaction mixture for expression in a mammalian cell, wherein the first and second nucleic acid sequences remain physically attached during the formatting.