Effect of tourniquet pressure and intra-individual variability on plasma fibrinogen, platelet P-selectin and monocyte tissue factor

Abstract

Small differences in levels of certain haemostatic components may be clinically significant. It is important therefore to eliminate potential sources of confounding variability. This study investigated the effect of removing tourniquet pressure prior to sample collection on plasma fibrinogen levels, platelet P-selectin and monocyte tissue factor expression. Blood was collected from the right arm under maintained tourniquet pressure and from the left arm following the release of pressure once the vein was sufficiently inflated for insertion of a needle. Whole blood was labelled within one hour of venepuncture to allow analysis of platelet P-selectin and monocyte tissue factor by flow cytometry. Plasma fibrinogen levels were analysed in samples stored at -70 degrees C, for all individuals at the end of the study using a method based on the Clauss technique. Intra-individual variability for each of the components was assessed by collecting samples under tourniquet pressure from four individuals on the same day on three consecutive weeks. Intra-individual variations were greater than assay CVs for all three components. There were no significant differences between the two tourniquet methods of collection for fibrinogen, P-selectin or tissue factor. In conclusion, there is no reason not to use a tourniquet during collection of blood for analysis of plasma fibrinogen, platelet P-selectin or monocyte tissue factor.

Preanalytical investigations of phlebotomy: methodological aspects, pitfalls and recommendations

“…Maybe, since laying in between clinics and laboratory, phlebotomy has long struggled to gain in scientific literature its own identity and the same reputation as laboratory assays. For instance, it’s symptomatic that just two out of the six papers issued by 1996-2000 used a kind of difference plot, and both of them neither mentioned the Bland-Altman eponymous nor cited the original paper (by the way, Scopus showed 3914 citations yet by that time) ( 19 , 20 ). Conversely, in the past years, mostly the last five (see Figure 1), we observed a change in trend and a growing attention payed toward this kind of research.…”

Venous Hypertension Induces Increased Platelet Reactivity and Accumulation in Patients with Chronic Venous Insufficiency

“…First, whole blood flow cytometric assays of platelet activation can minimize artifactual stimulation of platelets during sample handing. 21,[30][31][32] Platelets were directly analyzed in their physiological milieu of whole blood (including red cells and white cells, both of which could affect platelet activation). Second, it is important to determine the most sensitive flow cytometric parameter for evaluating platelet activation status.…”

Effect of nitric oxide modulation on systemic haemodynamics and platelet activation determined by P‐selectin expression

“…It is possible that discrepancies in results may be partly related to different protocols, experimental conditions and issues related to sample handling and analysis. We have previously spent a considerable amount of time optimizing the flow cytometry technique for the determination of P‐selectin expression on platelets (Ritchie et al , 1999, 2000a,b). This molecule is very easily upregulated during experimental procedures but great care was taken in this study to minimize artefacts that may confound interpretation of the results.…”

“…P‐selectin expression on platelets, PDMs and PLAs using flow cytometry. We have previously reported a method for P‐selectin expression on platelets (Ritchie et al , 1999, 2000a,b). Briefly, whole blood, 1 h after venepuncture, was diluted 1 in 8 with phosphate‐buffered saline (PBS) and incubated with phycoerythrin‐conjugated CD62P antibody (anti‐CD62P‐PE) [Becton Dickinson (BD) Cowley, Oxford, UK] and fluorescein isothiocyanate‐conjugated CD61 antibody (anti‐GPIIIa‐FITC) (BD) for 15 min at room temperature.…”