Validation of the Miceli Impedance Aggregometer With Donor Whole Blood: Testing Platelet Aggregation Using TRAP-6 Lyospheres and Varying Stir Speed

Abstract

Platelet function is critically examined for its role within cardiovascular disease, which is the leading cause of death in America, with over 650,000 per year. Patients with a medical history and risk factors derived from myocardial infarction, coronary artery disease, pulmonary embolism, arrhythmias, and valvular disease are often placed on therapies to reduce their risk of mortality. These therapies often include antiplatelet measures due to platelets being primary regulators in hemostasis and their involvement in atherosclerotic events by way of adhesion, activation, and aggregation. Impedance aggregometry is a lab-based test used to measure platelet aggregation and characterize the efficacy of antiplatelet intervention. The MICELI (MICrofluidic ELectrical Impedance) device is a scaled-down impedance aggregometer capable of measuring platelet aggregation as it occurs in small volume blood and is induced by various biochemical agonists, i.e. collagen, ADP, TRAP-6. The Protease-Activating Receptor-1 (PAR-1) in the cardiovascular system participates in blood clotting through thrombin signaling and is acted on by thrombin receptor-activating peptide-6 (TRAP-6). The purpose of this study is to improve the precision and ease of use of the prototype MICELI aggregometer by investigating the possibility of using pre-quantified TRAP-6 Lyospheres and finding the ideal RPM stir speed for optimal platelet aggregation. The efficacy of multiple TRAP-6 excipient types will be evaluated in this experiment, so the possibility of using them for point of care testing with a MICELI prototype in future studies may be determined. 7 The working hypotheses are: 1. The stir plate indicator setting of “9” for stir speed will produce uniform, high-amplitude platelet aggregation with minimal lag time (30 seconds) when TRAP-6 is used to induce aggregation in donor whole blood. Stir speed “9” was selected based off previous MICELI experiments that utilized RPM but were not focused on evaluating the changes in speeds. In those experiments higher speeds were utilized like “8” and “9”, while lower speeds and “10” were not. 2. The Lyophilized TRAP6 Excipient Types B469, B457, and B459 will produce statistically comparable or equal aggregation amplitude, area under the curve and lag time results to BioLyph Solvent TRAP-6 Agonist. Previous experiments utilizing ADP excipients, lyospheres, and solutions have been conducted that produced similar results to one another when their materials came from the same outsourced laboratories. This study will aim to utilize microfluidic electrical impedance for determining the optimal RPM setting to induce platelet aggregation while using TRAP-6 agonists. Secondly, the study will aim to evaluate efficacy of lyophilization of TRAP-6 utilizing BioLyph Solvent and multiple TRAP-6 excipient types (“B459”, “B457”, “B469”).