One-Step Recovery of scFv Clones from High-Throughput Sequencing-Based Screening of Phage Display Libraries Challenged to Cells Expressing Native Claudin-1

Figure 3

Evaluation of binding for scFv phages and IgG4. (a) The panel shows the binding of scFv phages, clones 3_2, 3_5, and 3_67, to HEK293 cells (gray bars) and to cells transduced with CLDN1 vector (HEK293-CLDN1, black bars). Clone 3_2 was discarded, because of nonspecific binding to HEK293-CLDN1 cells. (b) SDS-PAGE analysis of IgG4, converted from the scFv clones 3_5 and 3_67, as indicated. Samples in lanes 1 and 3 were run under nonreducing conditions, so that the whole IgG4 and the heavy-chain/light chain IgG4 dimers were accordingly visualized. Under reducing conditions, the IgG4 preparations showed the fully denatured light and heavy chains, as indicated. (c) The panels show the binding of the IgG4s, converted from scFv fragments 3_5 and 3_67, to HEK293 (gray lines) and to HEK293-CLDN1 (black lines) cells at increasing antibody concentrations.

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