Figure 4.

(A) Enrichment of the scFvhag ribosome complexes from mixtures with scFvAL2 by ribosome display. mRNA of scFvhag c5 was diluted 10⁸ times with mRNA of scFvAL2, and the mixture was used for ribosome display. After affinity selection of scFvhag ribosome complexes, mRNA was isolated and reverse transcribed to cDNA using oligonucleotide ON5, and cDNA was amplified by PCR using oligonucleotides ON3 and ON7 and analyzed by agarose electrophoresis. Lanes 1–5 are PCR products amplified after the first to fifth cycles of the ribosome display, respectively. Lane M is the 1-kb DNA ladder (GIBCO/BRL) used as molecular weight markers. PCR products corresponding to scFvhag and scFvAL2 are labeled. (B) Enrichment of either scFvhag c5 or scFvAL2 ribosome complexes by ribosome display as a function of immobilized antigen. mRNAs of scFvhag and scFvAL2 were mixed in a 1:1 ratio and used for one cycle of ribosome display. After affinity selection, mRNA was isolated, reverse transcribed, amplified by PCR, and analyzed by agarose electrophoresis as in A. The same 1:1 mixture was affinity selected on immobilized transferrin (tr) as a control, ampicillin-transferrin conjugate (amp), or hemagglutinin-peptide-transferrin conjugate (hag). PCR products corresponding to scFvhag and scFvAL2 are labeled.