THE IMPORTANCE OF PLATELETS IN THE EXPRESSION OF MONOCYTE TISSUE FACTOR ANTIGEN MEASURED BY A NEW WHOLE-BLOOD FLOW CYTOMETRIC ASSAY

Previous methods for the determination of monocyte tissue factor (TF) have been technically complex, difficult to standardize, prone to spur iously elevated results and difficult to implement in a clinical labor atory environment. We report the development of a two-color whole bloo d cytometric technique that overcomes many of these disadvantages. The assay uses small volumes of citrated blood (1.0 mi), can be performed in under one hour (if endotoxin stimulation is not per; formed), is r eproducible (CV = 5%) and uses methodology commonly available in clini cal laboratories. Baseline (mean +/- SD) expression of monocyte TF in normal subjects was very low (1.1 +/- 0.95%, Mean Fluorescence [Mean F L] 0.20 +/- 0.01) making relatively small increases easy to detect. Mo nocyte TF expression following endotoxin (LPS) stimulation for 1 h was 34.6 +/- 11.2% (Mean FL 0.32 +/- 0.04). LPS-stimulated activity varie d between subjects (21-68%) but was remarkably consistent for individu al subjects (CV = 5.4%). Stimulated monocyte TF expression was directl y proportional to the platelet count and was reduced by platelet prote ctive anticoagulants and by ingestion of aspirin. Non LPS-stimulated m onocyte TF was markedly increased, in a dose-dependent manner, by addi ng collagen to whole blood. This was apparently associated with platel et-monocyte binding and could be abolished by anti-P-Selectin. We conc lude that the whole blood flow cytometric assay of monocyte TF may be a valuable tool for clinical use and a useful model system for evaluat ing the humoral and cellular factors governing monocyte TF expression in a natural environment.