Detection of MPL exon 10 mutations. (A) Sequence traces of the 4 MPL exon 10 mutant alleles, showing 2 different mutations leading to the same MPL W515K substitution (MPL W515Ki and MPL W515Kii), and MPL S505N as an acquired mutation. (B) Allele-specific PCR strategy showing the common forward and reverse intronic primers and the allele-specific primer within MPL exon 10. (C) Mixing experiments with normal and mutant DNA demonstrating the sensitivity of the allele-specific PCR assays used for mutation screening. The images were captured on a Gel Doc 200 imager (Bio-Rad, Hercules, CA) using Quantity One software (Bio-Rad). NA indicates T cell sample not available from this patient; Pt, patient sample; WT, wild-type; NTC, no template control.