Purifying platelets with high-throughput microfluidics for pediatric and neonatal transfusions - Platelet (PLT) transfusions are necessary to prevent and treat bleeding. However, children and neonates have worse outcomes with platelet transfusions. Despite being lifesaving, PLT transfusions may lead to transfusion-related complications, such as transfusion related acute lung injury, transfusion associated circulatory overload, allergic reactions, and febrile non- hemolytic transfusion reactions. These complications are largely due to the contaminants that accumulate in the PLT suspending medium. Common methods of PLT processing include centrifugation and leukoreduction filters. However, each of these modalities have significant drawbacks, including premature PLT activation, creation of inflammatory microparticles, and inefficiency to remove all contaminants (whether large or small). Our objective is to develop and validate a simple-to-use, disposable device capable of removing all contaminants while preserving the highest-functioning PLTs. Our novel microfluidic technology, controlled incremental filtration (CIF), separates blood cells by size using devices with compact footprints and minimal void volumes, while minimizing cell activation and ensuring practical flow rates. Our latest CIF device prototypes remove ≥60% of supernatant and recover 98% of large, highly functional PLT (~90% overall PLT recovery), while minimally increasing PLT activation. The purpose of this study is to test our hypothesis that purifying PLTs with high-throughput CIF- based microfluidics is feasible, effective, and safe. We will test this hypothesis through three Specific Aims. In Aim 1, we will optimize the efficiency of the CIF devices and scale them up to be able to run at clinically relevant flow rates using human PLT units obtained from a regional blood blank. In Aim 2 we will evaluate the effect of CIF purification on the hemostatic function of PLTs in vitro (using a large battery of tests, including flow cytometry, light transmission aggregometry, and flow adhesion assays (type I collagen, VWF, and thrombin) and in vivo by transfusing purified human PLT into thrombocytopenic NOD/SCID mice and evaluating thrombus formation time using photochemical and ferric chloride injury models. In Aim 3, we will evaluate the safety and efficacy of PLTs purified by CIF in a premature neonatal pig model, including piglets with necrotizing enterocolitis (NEC). We hypothesize that transfusion of CIF- processed pig PLT into premature neonatal pigs will be safer (less inflammation) and more efficacious (less bleeding) than conventional PLT processing methods in piglets with and without NEC. This project establishes the essential foundation for finalizing the device design and conducting pivotal human trials, paving the way for safer PLT transfusions in all patients.
