Individual Platelet Adhesion (iPA): A novel assay of platelet function
Creators: name: Alonso, Ana Lopez
Publisher: name: Royal College of Surgeons in Ireland
Year: 2019
Subjects: Uncategorized

description: Cardiovascular disease is the leading cause of mortality in the developed countries and platelet related thrombosis the main complication in this condition. Anti-platelet agents are commonly used in the prevention of acute cardiovascular events. According to the American Heart Association and the American College of Cardiology (Circulation 2012), a class IIb recommendation is given for the use of platelet function test. This approach has been implemented in several clinical trials in order to improve outcomes in cardiovascular patients [1]. There are a number of platelet function tests available for clinical use but there is a lack of correlation between the different tests and none of them are routinely used to assess anti-platelet therapy effect. We report a rapid and accurate methodology to quantify platelet-protein interactions termed <em>iPA</em>. This novel technique contains a micro-array of 6-μm fibrinogen dots which can capture platelets from whole blood, one platelet per dot. The fractional occupancy of an array of fibrinogen dots after a predefined incubation time (specifically 30 minutes) quantitatively assays platelet adhesion to the protein matrix. We demonstrated that <em>iPA</em> enables detection of the effects of aspirin, P2Y<sub>12 </sub>inhibitors and αIIbβ3 receptor inhibitors <em>in vitro</em>. The mechanism of <em>iPA</em> to detect the effectiveness of anti-platelet therapy is based on a decrease of platelet adhesion upon addition of an agonist. An important factor to note regarding <em>iPA</em> is that in comparison to a fibrinogen coated array, the profile of platelet adhesion is different. On a static fibrinogen coated array platelet agonists would normally increase adhesion and spreading due to an increase in platelet activation. However the same is not true for <em>iPA</em>. Platelet activation induces aggregation and the aggregates thus are unable to bind during the assay. The reasoning behind these events is that firstly <em>iPA</em> is performed on a rocker, thus not allowing platelet aggregates to adhere gravitationally. Secondly the size of the dot is optimized to capture a single platelet. This agonist induced decrease in adhesion is inhibited by the specific anti-platelet drug and this is in fact the mechanism of detection. To summarise, in <em>iPA </em>normal resting platelets adhere maximally. Platelets that are in the presence of a platelet inhibitor also adhere normally to the 6-μm fibrinogen dots. Platelets stimulated with agonists show a marked reduction in fibrinogen occupancy. Platelets in the presence of antagonists and agonists show an occupancy value that is dependent on the effectiveness of the anti-platelet agent over the agonist. Furthermore we showed that <em>iPA</em> can specifically detect the effect of each drug in patients treated with these anti-platelet agents. <em>iPA</em> correlated well with the standard techniques of flow cytometry and light transmission aggregometry. Furthermore, <em>iPA</em> can be easily integrated with microfluidic platforms. Overall it promises to be more effective and rapid in the assessment of drug effects of anti-platelet agents during the treatment of cardiovascular disease. <strong> <br></strong> <strong><br> </strong>