A high-throughput microfluidic approach for 1000-fold leukocyte reduction of platelet-rich plasma

Leukocyte reduction of donated blood products substantially reduces the risk of a number of transfusion-related complications. Current ‘leukoreduction’ filters operate by trapping leukocytes within specialized filtration material, while allowing desired blood components to pass through. However, the continuous release of inflammatory cytokines from the retained leukocytes, as well as the potential for platelet activation and clogging, are significant drawbacks of conventional ‘dead end’ filtration. To address these limitations, here we demonstrate our newly-developed ‘controlled incremental filtration’ (CIF) approach to perform high-throughput microfluidic removal of leukocytes from platelet-rich plasma (PRP) in a continuous flow regime. Leukocytes are separated from platelets within the PRP by progressively syphoning clarified PRP away from the concentrated leukocyte flowstream. Filtrate PRP collected from an optimally-designed CIF device typically showed a ~1000-fold (i.e. 99.9%) reduction in leukocyte concentration, while recovering >80% of the original platelets, at volumetric throughputs of ~1 mL/min. These results suggest that the CIF approach will enable users in many fields to now apply the advantages of microfluidic devices to particle separation, even for applications requiring macroscale flowrates.

[1]  P. Holme,et al.  Shear-induced platelet activation and platelet microparticle formation in native human blood. , 1998, Thrombosis research.

[2]  D. Devine,et al.  Effects of prestorage white cell reduction on platelet aggregate formation and the activation state of platelets and plasma enzyme systems , 1999, Transfusion.

[3]  G. Burns,et al.  GMP-140 binding to neutrophils is inhibited by sulfated glycans. , 1991, The Journal of biological chemistry.

[4]  H. Carper,et al.  In vitro evaluation of a high-efficiency leukocyte adherence filter , 1991, Annals of Hematology.

[5]  S. Dzik Leukodepletion blood filters: filter design and mechanisms of leukocyte removal. , 1993, Transfusion medicine reviews.

[6]  Robert H. Austin,et al.  Deterministic separation of cancer cells from blood at 10 mL/min , 2012 .

[7]  C. W. Heegaard,et al.  Measurement of phosphatidylserine exposure during storage of platelet concentrates using the novel probe lactadherin: a comparison study with annexin V , 2009, Transfusion.

[8]  Aram J. Chung,et al.  Continuous inertial microparticle and blood cell separation in straight channels with local microstructures. , 2016, Lab on a chip.

[9]  Ian Papautsky,et al.  Continuous separation of blood cells in spiral microfluidic devices. , 2013, Biomicrofluidics.

[10]  V. Vandelinder,et al.  Separation of plasma from whole human blood in a continuous cross-flow in a molded microfluidic device. , 2006, Analytical chemistry.

[11]  Sergey S Shevkoplyas,et al.  Biomimetic autoseparation of leukocytes from whole blood in a microfluidic device. , 2005, Analytical chemistry.

[12]  D. Inglis,et al.  A scalable approach for high throughput branch flow filtration. , 2013, Lab on a chip.

[13]  S. McCluskey,et al.  Red cell transfusion and the immune system , 2015, Anaesthesia.

[14]  V. Vicente,et al.  Evaluation of Leukocyte–Depleted Platelet Concentrates Obtained by In–Line Filtration , 2000, Vox Sanguinis.

[15]  M. Yamada,et al.  Hydrodynamic filtration for on-chip particle concentration and classification utilizing microfluidics. , 2005, Lab on a chip.

[16]  R. Tompkins,et al.  A microfluidics approach for the isolation of nucleated red blood cells (NRBCs) from the peripheral blood of pregnant women , 2008, Prenatal diagnosis.

[17]  Han Wei Hou,et al.  Microfluidic Devices for Blood Fractionation , 2011, Micromachines.

[18]  N. Blumberg,et al.  Optimizing platelet transfusion therapy. , 2004, Blood reviews.

[19]  Robert E. Nordon,et al.  Scaling deterministic lateral displacement arrays for high throughput and dilution-free enrichment of leukocytes , 2011 .

[20]  M. Wadhwa,et al.  Cytokine levels as performance indicators for white blood cell reduction of platelet concentrates , 2002, Vox sanguinis.

[21]  R S Reneman,et al.  Wall shear rate in arterioles in vivo: least estimates from platelet velocity profiles. , 1988, The American journal of physiology.

[22]  Sergey S Shevkoplyas,et al.  Controlled incremental filtration: a simplified approach to design and fabrication of high-throughput microfluidic devices for selective enrichment of particles. , 2014, Lab on a chip.

[23]  N. Blumberg,et al.  An association between decreased cardiopulmonary complications (transfusion‐related acute lung injury and transfusion‐associated circulatory overload) and implementation of universal leukoreduction of blood transfusions , 2010, Transfusion.

[24]  A. B. Frazier,et al.  Lateral-driven continuous dielectrophoretic microseparators for blood cells suspended in a highly conductive medium. , 2008, Lab on a chip.

[25]  B. Vanhaesebroeck,et al.  Platelet PI3Kβ and GSK3 regulate thrombus stability at a high shear rate. , 2015, Blood.

[26]  Murat Karabacak,et al.  Continuous Flow Microfluidic Bioparticle Concentrator , 2015, Scientific Reports.

[27]  Siyang Zheng,et al.  Streamline-Based Microfluidic Devices for Erythrocytes and Leukocytes Separation , 2008, Journal of microelectromechanical systems.

[28]  Xing Chen,et al.  Microfluidic chip for blood cell separation and collection based on crossflow filtration , 2008 .

[29]  N. Blumberg,et al.  Circulating immune complexes involving the ABO system after platelet transfusion , 1993, British journal of haematology.

[30]  J. O'brien,et al.  Shear stress activation of platelet glycoprotein IIb/IIIa plus von Willebrand factor causes aggregation: filter blockage and the long bleeding time in von Willebrand's disease , 1987 .

[31]  Mehmet Toner,et al.  Microfluidic diffusive filter for apheresis (leukapheresis). , 2006, Lab on a chip.

[32]  Wolfgang Schramm,et al.  Mechanism of platelet adhesion to von Willebrand factor and microparticle formation under high shear stress. , 2006, Blood.

[33]  G. Schoch,et al.  A comparison of filtered leukocyte-reduced and cytomegalovirus (CMV) seronegative blood products for the prevention of transfusion-associated CMV infection after marrow transplant. , 1995, Blood.

[34]  Roland Zengerle,et al.  Leukocyte enrichment based on a modified pinched flow fractionation approach , 2013 .

[35]  N. Marwaha,et al.  Leukoreduced blood components: Advantages and strategies for its implementation in developing countries , 2010, Asian journal of transfusion science.

[36]  Eugene J. Lim,et al.  Microfluidic, marker-free isolation of circulating tumor cells from blood samples , 2014, Nature Protocols.

[37]  J. Feijen,et al.  The mechanisms of leukocyte removal by filtration. , 1995, Transfusion medicine reviews.

[38]  Edward L Snyder,et al.  Reduction of febrile but not allergic reactions to RBCs and platelets after conversion to universal prestorage leukoreduction , 2004, Transfusion.

[39]  J. Hoxie,et al.  Detection of activated platelets in whole blood using activation- dependent monoclonal antibodies and flow cytometry , 1987 .

[40]  Sriram Neelamegham,et al.  von Willebrand factor self-association on platelet GpIbalpha under hydrodynamic shear: effect on shear-induced platelet activation. , 2010, Blood.

[41]  K S Sakariassen,et al.  Shear-induced platelet activation and platelet microparticle formation at blood flow conditions as in arteries with a severe stenosis. , 1997, Arteriosclerosis, thrombosis, and vascular biology.

[42]  J. Sturm,et al.  Continuous Particle Separation Through Deterministic Lateral Displacement , 2004, Science.