Analysis of clonotype specificity of cultured PBMC from patient 1. (A) Fluorescent spectratype. The nature of the sample analyzed is identified above each lane. The first 4 lanes are from 14-day cultures containing no addition (media), media alone, heparin alone (heparin), PF4 alone (PF4), or heparin:PF4 complexes (Hep/PF4). The next lane is from uncultured PBMC. The next 2 lanes are from the plasmid DNA clones used to generate the clonotype sequences. The lane marked “PCR blank” is a water control. The actin control amplification is shown as a separate subpanel. (B) Hybridization of DNA from the gel shown in A with a probe for clonotype 1b (radioautogram). The gel was electroblotted onto a nylon membrane and treated as described in Materials and Methods. (C) Rehybridization of the electroblot with a probe specific for clonotype 1a. The membrane was stripped of the previous probe and rehybridized as described. (D) Rehybridization of the membrane with a TCR β constant region probe after stripping of the previous probe.