Effect of heparanase inhibitor on tissue factor overexpression in platelets and endothelial cells induced by anti‐β2‐GPI antibodies: Comment from Mackman et al.

We read the article entitled “Effect of heparanase inhibitor on tissue factor overexpression in platelets and endothelial cells induced by anti-β2-GPI antibodies” by Capozzi et al. in the recent issue of Journal of Thrombosis and Haemeostasis. In this article, the authors used western blotting to measure levels of tissue factor (TF) protein in human endothelial cells and platelets. TF was detected using a rabbit anti-TF monoclonal antibody (mAb) from Abcam and was labeled as a narrow 33-kDa band in both endothelial cells and platelets (Figure 6). No molecular weight standards were shown in the figure.  Unfortunately, the catalog number of the antibody was not listed in the methods. Abcam sells six different rabbit anti-TF mAbs (catalog numbers: ab228968, ab 211016, ab252918, ab254010, ab242432, and ab254007). The datasheets state that the antibodies detect a band of approximately either 42 or 47 kDa (with a predicted molecular weight of 33 kDa). In addition, Abcam used to sell another rabbit anti-TFmAb, ab151748. However, the company found that ab151748 did not detect TF but instead bound to a nonspecific band and stopped selling it (https://www.abcam.com/tissu e-factor-antibody-epr8986-ab151748.html). Unfortunately, 29 papers have used this antibody to measure TF in human samples and other species, including 15 papers that used the antibody for western blotting. In a previous paper, Capozzi et al. used western blotting to measure TF protein expression in platelets. As with the Journal of Thrombosis and Haemostasis publication, the TF band was labeled as 33 kDa (Figure 1C). The source of the anti-TF monoclonal was listed as NF-κB Millipore, Darmstadt, Germany, which does not make sense. Notably, TF expression by platelets is controversial and therefore it is very important to validate the reagents used to detect TF in these cells.