Immunological characterization of scFvE2/p17: epitope specificity and affinity. (a), Soluble scFvE2/p17 from Sf9 cell lysates was analyzed in competition ELISA using solid support-adsorbed p17.1 versus soluble p17.1 (homologous, positive control), or versus MAp17 epitope variants p17.3, p17.7 and p17.9. Negative controls consisted of ELISA in the absence of competing peptide (no competitor), or ELISA in the presence of irrelevant antigen (CD147), or irrelevant scFv (scFv-M6-1B9). Average of three separate experiments, m ± SEM. (b), Isolation of H6MA-CA-scFvE2/p17 complexes. Lysates of cells coexpressing H6MA-CA and HA-tagged scFvE2/p17 were separated on affinity matrix consisting of anti-HA tag antibody-coupled agarose beads, and fractions analyzed by SDS-PAGE and Western blotting, using anti- His6 tag and anti-HA tag antibodies. Lane 1, whole cell lysate; lane 2, flow-through fraction; lane 3, column wash; lane 4, fraction eluted with SDS-PAGE loading buffer.