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Sorting of the library for high affinity clones. An FCHO cell pool stably transfected with mutant library was double-labeled using both PE-conjugated anti-Lambda chain antibody and FITC-conjugated OX40 ligand (100 nM) and analyzed by flow cytometry. Cells located in gate R3 (0.1%) were single-cell sorted into a 96-well plate with one cell per well and expanded for further analysis.
