The primary focus of our recent report1 is to examine whether Cys186-Cys209 disulfide–mutated tissue factor (TF) mimics cryptic TF and the importance of Cys186-Cys209 disulfide bond formation in TF decryption, 2 key issues that were not examined earlier. 2 In our view, one should be careful not to equate TF mutants that exhibit low procoagulant activity with cryptic TF. This incorrectly broadens the definition of cryptic TF and creates unnecessary confusion and controversy. The classic definition of cryptic TF is that it can form a stable complex with factor VIIa (FVIIa), but the resultant TF-FVIIa complexes fail to activate its macromolecular substrates, factors IX and X. 3, 4 Therefore, when probing the cryptic nature of TF, one should evaluate the activity of TF-FVIIa complexes at the cell surface. Despite some inherent differences in their binding properties, both cryptic and decrypted TF form stable high-affinity associations with FVII/FVIIa, well below plasma concentrations of FVII (see references in Bach and Monroe5). Cryptic TF would remain cryptic even in the presence of supraphysiologic concentrations of FVIIa. TF mutants that exhibit lower procoagulant activity at physiologic concentrations of FVII/FVIIa, but could be rescued by supraphysiologic concentrations of FVIIa should not be equated with cryptic TF. The Cys186-Cys209 disulfide–mutated TF falls into this category. They exhibit low procoagulant activity because the Cys186-Cys209 disulfide bond is required for efficient FVIIa binding1, 2 and not because they conform to cryptic TF form.

One of the criticisms of our study raised by Ruf and Versteeg is that in concluding that the allosteric disulfide bond is not essential for TF’s procoagulant activity, we took into consideration only data at supraphysiologic concentrations of TF ligands. Because the physiologic concentration of FVIIa was not sufficient to saturate the TF mutants at the cell surface, we had to use higher concentrations of FVIIa. We actually measured the amount of TF-FVIIa complexes formed at the cell surface and used this number to determine the specific activities of wild-type and TF mutants. This