Functional tests of ADAMTS13 variant expression secretion and activity. (A) Cell lines expressing wild-type, Q448E, and A732V ADAMTS13 were generated by using an FLP-recombinase system in 293 cells. Q448E is rs2301612 and A732V is rs41314453. (B-C) Concentrations of ADAMTS13 in (B) conditioned media and (C) cell lysate were measured by using AlphaLISA and adjusted by cell lysate glyceraldehyde-3-phosphate dehydrogenase levels to account for variations in cell number. Mean and standard deviations are plotted as well as individual values. (D) Recombinant enzyme-specific activity was measured with FRETS-VWF73 substrate. Differences in group levels were determined by the Mann Whitney U test. 1-8, thrombospondin type 1 repeats; CUB, C1r-C1s urinary epidermal growth factor bone morphogenetic protein domains; Cys-rich, cysteine-rich domain; P, propeptide; S, signal peptide.