Specific peptide inhibition of phage particle binding to anti-Ap^1-17 IgG demonstrates that phage insert sequences recognize the antigen-combining sites of anti-Ap^1-17 IgG.

<p>Microtiter plates were coated with affinity-purified anti-Ap^1-17 IgG from pt4 (A.) or pt14 (B.). After the blocking of free protein-binding sites with PBS-BSA, wells were incubated for 2 h with 50 µl PBS containing serial dilutions of either free (closed symbols) or KLH-conjugated (open symbols) Ap^1-17 (rhombus), Ap^17-30 (triangle), CBp^1-13 (circle) or Qp-1a (square). Then, without removing the inhibitor, 50 µl/well of an appropriate dilution of phage supernatant pc4.33 (A.) and pc14.49 (B.) was added and incubation prolonged for 2 h. Bound phage particles were detected by sequential addition of HRP-anti-M13 mAb and o-phenylenediamine solution. Results are expressed as percentage of binding inhibition. Each data point is the mean of duplicate wells. The data are representative of two experiments. The maximum amount of inhibitor (800 µg) corresponds to 382 nmol Ap^1-17, 558 nmol Ap^17-30, 486 nmol CBp^1-13 and 574 nmol Qp-1a.