Differential ability of tissue factor antibody clones on detection of tissue factor in blood cells and microparticles.
BACKGROUND
Tissue factor (TF), the primary initiator of coagulation in vivo, plays a major role in both thrombosis and hemostasis. The expression of TF in monocytes is well documented, but its presence in other blood cells has been disputed, possibly due to methodological variations among different studies.


METHODS
We studied TF expression on platelets, monocytes, lymphocytes and microparticles (MPs) by flow cytometry (FCM) with five commercially available mouse anti-human TF antibodies (HTF-1, TF9-10H10, CLB/TF-5, VIC7 and VD8). The ability of different TF antibodies to inhibit cell surface TF activity was explored by incubating LPS-stimulated monocytes and MPs derived from LPS-stimulated monocytes (MMPs) with TF antibodies followed by measuring TF activity.


RESULTS
HTF-1 detected TF only on LPS-stimulated monocytes, whereas, TF9-10H10 and VD8 detected TF associated with MPs and MMPs in addition to LPS stimulated monocytes. Surprisingly, CLB/TF-5 and VIC7 detected TF on platelets, monocytes even under unstimulated conditions, in addition to MPs and MMPs. CLB/TF-5 also detected TF on unstimulated lymphocytes. Inhibitory studies showed that at a final concentration of 10 μg/mL, HTF-1, CLB/TF-5 and VD8 inhibited monocyte TF activity by 81-84% and MMP TF activity by 92-96%; whereas TF9-10H10 had no inhibitory effect on TF activity in monocytes and MMPs.


CONCLUSIONS
Our results suggest non-specific binding by the CLB/TF-5 and VIC7 antibodies in a FCM test system and explain at least some of the reports on TF presence in blood cells, particularly TF associated with platelets and MPs. TF9-10H10 and VD8 are more suitable to detect TF on MPs by FCM.
Hematological Research Group, Department of Clinical Medicine, University of Tromsø, Tromsø, Norway. manjunath.goolyam@uit.no
Manjunath Goolyam
MG
Basavaraj
Jan Ole
JO
Olsen
Bjarne
B
Østerud
John-Bjarne
JB
Hansen
D000328:Adult; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001773:Blood Cells / Q000737:chemistry* / Q000276:immunology*; D055252:Cell-Derived Microparticles / Q000737:chemistry* / Q000276:immunology*; D002478:Cells, Cultured; D005260:Female; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D008297:Male; D051379:Mice; D013925:Thromboplastin / Q000032:analysis*
D000911:Antibodies, Monoclonal; D013925:Thromboplastin