A new method for quantitative analysis of whole blood platelet interaction with extracellular matrix under flow conditions.
A new method and device in which whole blood platelet deposition and aggregation on extracellular matrix (ECM) under defined shear conditions is quantitatively evaluated was developed. A 0.25 mL aliquot of citrated whole blood is placed on ECM and a defined shear rate is applied for 2 min using a cone and plate device. This is followed by staining and measuring the number of stained objects, the percentage of ECM surface covered with stained objects and the average size of the objects using an image analyzer. When normal blood is analyzed, platelet deposition is a shear and a time dependent process, reaching maximal levels within 2 min at high shear rate (1300 s-1) of about 20% surface coverage and average aggregate size of about 40-50 microns 2. These two parameters demonstrated positive correlation with the platelet count and the hematocrit. Studies using samples from patients with von Willebrand disease (vWD) and Glanzmann's Thrombasthenia (GT) were performed and demonstrated the ability of the new method to detect these pathological conditions. Blood samples of vWD patients showed a very low adhesion and aggregation at high shear rate as reflected by very low surface coverage (5.2%) and average particle size of single platelets (21.3 microns2). GT samples at a high shear rate demonstrated surface coverage similar to normal blood samples (21.7%) but with average particle size of single platelets (21.3 microns2). The new method is an alternative method to clinically evaluate platelet function under close to physiological conditions.
Varon
D
D
National Hemophilia Center, Sheba Medical Center, Tel-Hashomer, Israel.
Dardik
R
R
Shenkman
B
B
Kotev-Emeth
S
S
Farzame
N
N
Tamarin
I
I
Savion
N
N
D001792:Blood Platelets / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism*; D006400:Hematocrit; D006405:Hematology / Q000379:methods; D006487:Hemostasis; D006801:Humans; D008855:Microscopy, Electron, Scanning; D010973:Platelet Adhesiveness; D010974:Platelet Aggregation; D010976:Platelet Count; D012212:Rheology; D013915:Thrombasthenia / Q000097:blood; D014842:von Willebrand Diseases / Q000097:blood