Figure 1.

Overview of the workflow used to generate the scFv libraries from B cells isolated from Balb/c and SJL mice. B cells are isolated from lymph nodes and encapsulated into droplets with oligo-dT beads and a lysis solution. mRNA-bound beads are purified from the droplets, and then injected into a second emulsion with an OE-RT-PCR amplification mix that generates DNA amplicons that encode scFv with native pairing of heavy and light chain Ig. Libraries of natively paired amplicons are then electroporated into yeast for scFv display. FACS is used to identify high affinity scFv. Finally, deep antibody sequencing is used to identify all clones in the pre- and post-sort scFv libraries.