Title: Development of a novel clinical platelet analyser
Funder: Department of Health and Social Care

Abstract: 1. Hardware development The hardware of the system is modelled upon a parallel plate flow chamber that is used in the Poole laboratory and illustrated in Appendix Fig. 1. Blood is passed over a variety of surfaces that mimic the structures that platelets encounter in a damaged blood vessel. In the simplest of these systems the surface is coated with collagen. The flow rate of the blood may be adjusted so as to mimic in-vivo arterial shear forces. In our research-based system, platelets are labelled with a fluorescent dye (DiOC6) and images of flowing blood and thrombus formation are captured as real time video. Sample images are shown in Appendix Fig. 2. The novelty of this system over available competitors is that it is much more physiological, as it mimics closely blood flow through a vessel. Hence it has a much greater potential to predict a thrombotic tendency or a bleeding disorder of platelet origin. The disadvantage of the current laboratory system, is that it requires a large and expensive microscope, and the analysis of data is long-winded and difficult. To overcome these issues we have designed and constructed a small, desktop imaging system using a scientific grade camera and dedicated optics with LED illumination that is portable, lightweight and robust. This instrument is computer controlled by our own software, which both collects images and generates numerical metrics from the images acquired using statistical algorithms novel to this application. We are requesting funding for 2 years for two staff for development of the system, which will involve: (i) miniaturization of the flow system and conversion to a blood drawing system, instead of syringe pump. We intend that a blood sample, in vacutainer tube (or ideally a finger prick sample), will be inserted into the machine and automatically fed into the analysis system at bedside. (ii) miniaturization, using microfluidics, of the parallel plate chamber, and development towards a disposable, single-use, cartridge-based system for the final clinical analyser. 2. Software Algorithm development This is the primary area of intellectual property associated with this development. We have developed a novel method of data analysis, where coefficients of platelet aggregation and adhesion are determined by statistical analysis of the images. The data are captured using our bespoke software, as shown in Appendix Fig. 3, and parameters of aggregation and adhesion calculated that may then be used to compare against reference ranges. Because a very large dataset is acquired there is the potential for deriving multiple other, more subtle, parameters of platelet function under flow in our system. The software will be developed to allow determination of these additional parameters and optimise the algorithm for sensitivity and specificity to certain clinical conditions, such as aspirin treatment response. 3. Clinical validation In our pilot studies (in Appendix Fig. 4) we have used 20 healthy volunteer donors, and have acquired flow-based thrombus formation data from blood either untreated, or treated with the clinically-used anti-platelet agents aspirin or abciximab. The latter is a potent blocker of platelet aggregation whereas aspirin has a more mild effect. The data from our new system show very clear delineation of platelet functional activities under these three conditions, and provide convincing evidence that the system will have capabilities of stratifying patient responsiveness to drugs, and is likely to be sensitive to changes in bleeding function and thrombotic potential. We will compare platelet function determined by the new rapid technology to that determined by light transmission aggregation. Preliminary clinical utility will be tested by estimating associations between test results from the new technology and bleeding/thrombotic clinical outcomes (post-operative blood loss, transfusion requirement, and thrombosis).

Lay abstract: The ability of blood to clot is critical to us to stop bleeding. There is however a wide variety of circumstances where the mechanisms underlying clot formation are abnormal or need to be controlled by drugs. People vary from those who bleed too easily through to those at risk of thrombosis, where blood clots too readily, blocking blood vessels and leading to conditions such as deep vein thrombosis or heart attack. Blood platelets are the central cell involved in this process. Although several bedside tests of how well platelets function are currently available for clinical use, none provide estimates accurate enough to predict whether a patient has a tendency to bleed or is at high risk of thrombosis. This is enormously important in a variety of hospital-based situations, including major surgery, patients on antithrombotic therapies and patients undergoing blood transfusions. We have therefore developed a new medical instrument for measuring platelet function. This instrument is readily developable into a bedside system with the potential to be transformational. The system will, when developed, take a single small blood sample and analyse platelet function within 3 minutes. Importantly our device will provide a numerical readout based upon a novel method of analysing the images of clots as they form under conditions much closer to those inside the patient s body than any instrument currently available. Our aim is to demonstrate the effectiveness of this novel tool in the clinic with a view to predicting clinical risks and to direct antiplatelet drug management on a patient-specific basis. Development will take place in 3 phases: (1) hardware refinement and miniaturisation, (2) software engineering and analytical tools development and (3) taking measurements from patients, such as those undergoing coronary artery bypass graft surgery, and comparing results with existing techniques and against clinical outcome.

Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES;4.2 EVALUATION OF MARKERS AND TECHNOLOGIES
Health categories: BLOOD;CARDIOVASCULAR