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Jarob Saker - One of the best experts on this subject based on the ideXlab platform.
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the use of a hematology analyzer with a new generation of software as an alternative to flow cytometry for enumerating residual white blood cells in blood components
Transfusion, 2020Co-Authors: Richard Alejo Blanco, Chloe Cavagnetto, Laura Willmott, Elif Aydogdu, Nicola Akinyemi, Helena Standring, Simon Procter, Stephen F Garner, Atsushi Shirakami, Jarob SakerAbstract:BACKGROUND: Leukoreduction of blood components was implemented to reduce transfusion-associated risks. The detection level for residual white blood cells (rWBCs) required to demonstrate Leukoreduction was originally considered too low for hematology analyzers. Developments enabling cell counts in body fluids have, however, renewed interest in rWBC counting. An assessment of Sysmex XN hematology analyzers with software offering automated rWBC enumeration intended for use on blood components was performed. STUDY DESIGN AND METHODS: Performance characteristics were determined using platelet, red blood cell (RBC), and plasma samples spiked with WBCs. Subsequently, components (platelets, n = 1367; and plasma, n = 80) were tested and results compared with flow cytometry, to monitor Leukoreduction efficiency to a level of less than 1 × 106 /unit. Components identified by flow cytometry as having poor Leukoreduction, exceeding this limit, were also tested (platelets, n = 3; and RBCs, n = 10). RESULTS: Linearity studies up to 32 WBCs/μL showed good correlation between observed and expected results (R2 > 0.9996). Precision analysis gave an average limit of quantitation of 2 WBCs/μL with coefficients of variation less than 20%. Average carryover was 0.1%. Plain sample tubes were a source of aberrant results with routine components. Using ethylenediaminetetraacetic acid tubes the analyzer gave results greater than 1 × 106 /unit in 2.7% of cases compared with 1.4% by flow cytometry, but overall results were within specification, with more than 90% of components having rWBC values below the limit. All incidences of poor Leukoreduction, with flow cytometry results greater than 13 rWBCs/μL were correctly identified, with an excellent correlation between results (R2 = 0.9818). CONCLUSION: The analyzer demonstrated acceptable performance characteristics for enumeration of rWBCs; consequently, additional multisite evaluations are warranted.
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the use of a hematology analyzer with a new generation of software as an alternative to flow cytometry for enumerating residual white blood cells in blood components
Transfusion, 2020Co-Authors: Richard Alejo Blanco, Chloe Cavagnetto, Laura Willmott, Elif Aydogdu, Nicola Akinyemi, Helena Standring, Simon Procter, Stephen F Garner, Atsushi Shirakami, Jarob SakerAbstract:Background Leukoreduction of blood components was implemented to reduce transfusion-associated risks. The detection level for residual white blood cells (rWBCs) required to demonstrate Leukoreduction was originally considered too low for hematology analyzers. Developments enabling cell counts in body fluids have, however, renewed interest in rWBC counting. An assessment of Sysmex XN hematology analyzers with software offering automated rWBC enumeration intended for use on blood components was performed. Study design and methods Performance characteristics were determined using platelet, red blood cell (RBC), and plasma samples spiked with WBCs. Subsequently, components (platelets, n = 1367; and plasma, n = 80) were tested and results compared with flow cytometry, to monitor Leukoreduction efficiency to a level of less than 1 × 106 /unit. Components identified by flow cytometry as having poor Leukoreduction, exceeding this limit, were also tested (platelets, n = 3; and RBCs, n = 10). Results Linearity studies up to 32 WBCs/μL showed good correlation between observed and expected results (R2 > 0.9996). Precision analysis gave an average limit of quantitation of 2 WBCs/μL with coefficients of variation less than 20%. Average carryover was 0.1%. Plain sample tubes were a source of aberrant results with routine components. Using ethylenediaminetetraacetic acid tubes the analyzer gave results greater than 1 × 106 /unit in 2.7% of cases compared with 1.4% by flow cytometry, but overall results were within specification, with more than 90% of components having rWBC values below the limit. All incidences of poor Leukoreduction, with flow cytometry results greater than 13 rWBCs/μL were correctly identified, with an excellent correlation between results (R2 = 0.9818). Conclusion The analyzer demonstrated acceptable performance characteristics for enumeration of rWBCs; consequently, additional multisite evaluations are warranted.
Christopher C. Silliman - One of the best experts on this subject based on the ideXlab platform.
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proteomic analysis of the supernatant of red blood cell units the effects of storage and leucoreduction
Vox Sanguinis, 2013Co-Authors: Monika Dzieciatkowska, Kevin J Land, Michael A Ellison, Marguerite R Kelher, Anirban Banerjee, Christopher C. Silliman, Ernest E Moore, F. Bernadette WestAbstract:Background Red blood cell (RBC) transfusion is a life-saving intervention for critically ill patients; however, it has been linked to increased morbidity and mortality. We hypothesize that a number of important proteins accumulate during routine storage of RBCs, which may explain some of the adverse effects seen in transfused patients. Study Design Five RBC units were drawn and divided (half prestorage leucoreduced (LR-RBC) and half left as an unmodified control (RBC). The supernatant was separated on days 1 and 42 of storage and proteomic analyses completed with in-gel tryptic digestion and nano-liquid chromatography tandem mass spectrometry. Results In RBC supernatants, 401 proteins were identified: 203 increased with storage, 114 decreased, and 84 were unchanged. In LR-RBC supernatant, 231 proteins were identified: 84 increased with storage, 30 decreased, and 117 were unchanged. Prestorage leucoreduction removed many platelet- and leucocyte-derived structural proteins; however, a number of intracellular proteins accumulated including peroxiredoxins (Prdx) 6 and latexin. The increases were confirmed by immunoblotting, including the T-phosphorylation of Prdx-6, indicating that it may be functioning as an active phospholipase. Active matrix metalloproteinase-9 also increased with a coinciding decrease in the metalloproteinase inhibitor 1 and cystatin C. Conclusion We conclude that a number of proteins increase with RBC storage, which is partially ameliorated with leucoreduction, and transfusion of stored RBCs may introduce mediators that result in adverse events in the transfused host.
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identification of lipids that accumulate during the routine storage of prestorage leukoreduced red blood cells and cause acute lung injury
Transfusion, 2011Co-Authors: Christopher C. Silliman, Marguerite R Kelher, Ernest E Moore, Samina Y Khan, Lauren Gellar, David J ElziAbstract:Transfusion-related acute lung injury (TRALI) remains the most common cause of transfusion-related mortality.1,2 TRALI has been linked to a number of mediators including antibodies against white blood cell (WBC) antigens, biologically active lipids that accumulate during routine storage of cellular blood components, and soluble CD40 ligand.3–6 During routine storage of red blood cells (RBCs), lipids accumulate in the plasma fraction, supernatant.7 These lipids consist of nonpolar lipids and a mixture of lysophosphatidylcholines (lyso-PCs), as defined by their retention times on normal-phase high-pressure liquid chromatography (HPLC).7 Lyso-PCs at concentrations that accumulate during RBC and platelet (PLT) concentrate storage precipitate polymorphonuclear neutrophil (PMN)-mediated endothelial damage and acute lung injury (ALI) in a two-event in vivo model of TRALI.8 Universal prestorage Leukoreduction has been instituted in a number of countries, and despite decreasing the numbers of febrile transfusion reactions, outside of a single institution in the United States, there has been no mention of its effects on the rates of TRALI.9,10 In addition, prestorage Leukoreduction by filtration is known to decrease WBC contamination by greater than 3 logs and decrease PLT contamination by 4 to 5 logs, as evidenced by eradication of soluble CD40 ligand accumulation during routine storage because it is PLT derived.3 It is hypothesized, therefore, that prestorage Leukoreduction changes the lipids that accumulate during routine storage but does not affect their ability to rapidly prime the PMN oxidase or to serve as the second event to precipitate ALI in an in vivo model.
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red blood cell supernatant potentiates lps induced proinflammatory cytokine response from peripheral blood mononuclear cells
Journal of Interferon and Cytokine Research, 2009Co-Authors: Joel M. Baumgartner, Trevor L Nydam, Jason H Clarke, Anirban Banerjee, Christopher C. Silliman, Martin D. MccarterAbstract:Allogeneic blood transfusion has an immunomodulatory capacity on its recipients through accumulation of immunologically active substances with blood storage, and prestorage Leukoreduction reduces many of these mediators. We investigated lipopolysaccharide (LPS)-induced cytokine response of peripheral blood mononuclear cells (PBMCs) exposed to packed red blood cell (PRBC) supernatants from leukoreduced (LR) or non-leukoreduced (NLR) units with variable duration of storage. PRBC units were collected with or without Leukoreduction on Day 0 before routine storage. The plasma fraction (supernatant) was isolated from LR and NLR units after 1 day (D1) or 42 days (D42) of storage and exposed to PBMCs versus control media for 24 h, then with LPS for an additional 24 h. Cell supernatants were analyzed for IL-1β, IL-6, IL-8, IL-10, and TNF-α by cytokine bead array. IL-1β, TNF-α, and IL-6 were significantly elevated in PRBC groups versus control. D42 NLR PRBC supernatant significantly increased secretion of IL-1β and IL-6 compared to D1 NLR PRBC supernatant. LR significantly attenuated the cytokine response of IL-1β. Thus, PRBC supernatant potentiates proinflammatory LPS-induced cytokine secretion from PBMCs. This response is accentuated with storage duration and partially attenuated with Leukoreduction. These findings may partially explain the immune activation seen clinically after blood transfusion.
Timothy A Pritts - One of the best experts on this subject based on the ideXlab platform.
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effects of whole blood Leukoreduction on platelet function and hemostatic parameters
Transfusion Medicine, 2019Co-Authors: Mackenzie C Morris, Philip C Spinella, Timothy A Pritts, Rosalie A Veile, Lou Ann Friend, W C Dorlac, Michael D GoodmanAbstract:Aims/objectives The aim of this study was to evaluate the hemostatic consequences of whole blood Leukoreduction (LR). Background Whole blood is being used for trauma resuscitation in the military, and an increasing number of civilian trauma centres across the nation. The benefits of LR, such as decreased infectious and transfusion-related complications, are well established, but the effects on hemostatic parameters remain a concern. Methods Twenty-four units of whole blood were assigned to one of the four groups: non-leukoreduced (NLR), leukoreduced at 1 h and a height of 33 in. (LR-1), leukoreduced at 4 h and a height of 33 in. (LR-4(33)), or leukoreduced at 4 h and a height of 28 in. (LR-4(28)). Viscoelastic parameters, platelet aggregation, cell counts, physiological parameters and thrombin potential were evaluated immediately before and after LR, and on days 1, 7, 14 and 21 following LR. Results The viscoelastic parameters and thrombin generation potential were unchanged between the groups. Platelet aggregation was reduced in the LR-1 group compared with NLR after 7 days. The LR-4(28) group also showed a trend of reduced platelet aggregation compared with NLR. Aggregation in LR-4(33) was similar to NLR throughout the storage time. Physiological and electrolyte changes over the whole blood storage period were not affected by LR. Conclusion Our study shows that whole blood can be LR at 4 h after collection and a height of 33 in. while maintaining platelet count and without altering platelet function and hemostatic performance.
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Leukoreduction of packed red blood cells attenuates proinflammatory properties of storage derived microvesicles
Journal of Surgical Research, 2018Co-Authors: Jillian R Richter, Jeffrey M Sutton, Phillip Hexley, Taylor A Johannigman, Alex B Lentsch, Timothy A PrittsAbstract:Abstract Background Leukoreduction prior to packed red blood cell (pRBC) storage is not a universally accepted practice. Our laboratory has previously shown that microvesicles (MVs) accumulate in pRBC units during storage and play an important role in lung injury after resuscitation. Currently, the effect of Leukoreduction on MV formation in stored pRBC units is unknown. In the present study, we investigated the hypothesis that Leukoreduction of pRBC units prior to storage would attenuate the production of MVs and decrease pulmonary inflammation after hemorrhage and resuscitation. Methods Leukoreduced and nonleukoreduced pRBC units were prepared from human donors and C57/Bl6 mice and stored for up to 42 d and 14 d, respectively. At intervals during storage, MVs were isolated from pRBC units, quantified and characterized based on size, morphology, and levels of proinflammatory cytokines. In additional experiments, mice underwent controlled hemorrhage followed by resuscitation with normal saline (NS) with or without equal numbers of MVs isolated from leukoreduced or nonleukoreduced stored mouse pRBC. Histologic lung sections were evaluated for the presence of tissue edema and inflammatory cells. Results For both human and mouse pRBCs, the number of MVs significantly increased throughout the storage period. There were significantly fewer MVs present in leukoreduced units. The average MV size significantly increased over time and was similar between groups. Levels of interleukin 1α (IL-1α), regulated on activation, normal T cell expressed and secreted (RANTES), and macrophage-derived chemokine (MDC) were lower in MVs from leukoreduced pRBC units as compared with MVs from nonleukoreduced units. Hemorrhaged mice resuscitated with NS with the addition of MV from leukoreduced pRBC demonstrated significantly less pulmonary edema and inflammatory cell recruitment as compared to those resuscitated with NS with the addition of MV from nonleukoreduced pRBC. Conclusions Prestorage Leukoreduction of pRBC units reduces the formation and proinflammatory properties of MV, which in turn decreases lung injury secondary to MV from stored pRBC units after hemorrhage and resuscitation.
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role of Leukoreduction of packed red blood cell units in trauma patients a review
International journal of hematology research, 2016Co-Authors: Young Kim, Brent T Xia, Alex L Chang, Timothy A PrittsAbstract:Hemorrhagic shock is a leading cause of mortality within the trauma population, and blood transfusion is the standard of care. Leukoreduction filters remove donor leukocytes prior to transfusion of blood products. While the benefits of leukocyte depletion are well documented in scientific literature, these benefits do not translate directly to the clinical setting. This review summarizes current research regarding Leukoreduction in the clinical arena, as well as studies performed exclusively in the trauma population.
Richard Alejo Blanco - One of the best experts on this subject based on the ideXlab platform.
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the use of a hematology analyzer with a new generation of software as an alternative to flow cytometry for enumerating residual white blood cells in blood components
Transfusion, 2020Co-Authors: Richard Alejo Blanco, Chloe Cavagnetto, Laura Willmott, Elif Aydogdu, Nicola Akinyemi, Helena Standring, Simon Procter, Stephen F Garner, Atsushi Shirakami, Jarob SakerAbstract:BACKGROUND: Leukoreduction of blood components was implemented to reduce transfusion-associated risks. The detection level for residual white blood cells (rWBCs) required to demonstrate Leukoreduction was originally considered too low for hematology analyzers. Developments enabling cell counts in body fluids have, however, renewed interest in rWBC counting. An assessment of Sysmex XN hematology analyzers with software offering automated rWBC enumeration intended for use on blood components was performed. STUDY DESIGN AND METHODS: Performance characteristics were determined using platelet, red blood cell (RBC), and plasma samples spiked with WBCs. Subsequently, components (platelets, n = 1367; and plasma, n = 80) were tested and results compared with flow cytometry, to monitor Leukoreduction efficiency to a level of less than 1 × 106 /unit. Components identified by flow cytometry as having poor Leukoreduction, exceeding this limit, were also tested (platelets, n = 3; and RBCs, n = 10). RESULTS: Linearity studies up to 32 WBCs/μL showed good correlation between observed and expected results (R2 > 0.9996). Precision analysis gave an average limit of quantitation of 2 WBCs/μL with coefficients of variation less than 20%. Average carryover was 0.1%. Plain sample tubes were a source of aberrant results with routine components. Using ethylenediaminetetraacetic acid tubes the analyzer gave results greater than 1 × 106 /unit in 2.7% of cases compared with 1.4% by flow cytometry, but overall results were within specification, with more than 90% of components having rWBC values below the limit. All incidences of poor Leukoreduction, with flow cytometry results greater than 13 rWBCs/μL were correctly identified, with an excellent correlation between results (R2 = 0.9818). CONCLUSION: The analyzer demonstrated acceptable performance characteristics for enumeration of rWBCs; consequently, additional multisite evaluations are warranted.
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the use of a hematology analyzer with a new generation of software as an alternative to flow cytometry for enumerating residual white blood cells in blood components
Transfusion, 2020Co-Authors: Richard Alejo Blanco, Chloe Cavagnetto, Laura Willmott, Elif Aydogdu, Nicola Akinyemi, Helena Standring, Simon Procter, Stephen F Garner, Atsushi Shirakami, Jarob SakerAbstract:Background Leukoreduction of blood components was implemented to reduce transfusion-associated risks. The detection level for residual white blood cells (rWBCs) required to demonstrate Leukoreduction was originally considered too low for hematology analyzers. Developments enabling cell counts in body fluids have, however, renewed interest in rWBC counting. An assessment of Sysmex XN hematology analyzers with software offering automated rWBC enumeration intended for use on blood components was performed. Study design and methods Performance characteristics were determined using platelet, red blood cell (RBC), and plasma samples spiked with WBCs. Subsequently, components (platelets, n = 1367; and plasma, n = 80) were tested and results compared with flow cytometry, to monitor Leukoreduction efficiency to a level of less than 1 × 106 /unit. Components identified by flow cytometry as having poor Leukoreduction, exceeding this limit, were also tested (platelets, n = 3; and RBCs, n = 10). Results Linearity studies up to 32 WBCs/μL showed good correlation between observed and expected results (R2 > 0.9996). Precision analysis gave an average limit of quantitation of 2 WBCs/μL with coefficients of variation less than 20%. Average carryover was 0.1%. Plain sample tubes were a source of aberrant results with routine components. Using ethylenediaminetetraacetic acid tubes the analyzer gave results greater than 1 × 106 /unit in 2.7% of cases compared with 1.4% by flow cytometry, but overall results were within specification, with more than 90% of components having rWBC values below the limit. All incidences of poor Leukoreduction, with flow cytometry results greater than 13 rWBCs/μL were correctly identified, with an excellent correlation between results (R2 = 0.9818). Conclusion The analyzer demonstrated acceptable performance characteristics for enumeration of rWBCs; consequently, additional multisite evaluations are warranted.
Darrell J Triulzi - One of the best experts on this subject based on the ideXlab platform.
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effects of storage and Leukoreduction on lymphocytes and epstein barr virus genomes in platelet concentrates
Transfusion, 2009Co-Authors: David T Rowe, Albert D Donnenberg, D L Griffin, Darrell J TriulziAbstract:BACKGROUND: Epstein-Barr virus (EBV) persists in infected B lymphocytes in blood donors. Lymphocytes are viable during platelet (PLT) storage. The effects of storage and Leukoreduction on lymphocytes and EBV genomes are evaluated. STUDY DESIGN AND METHODS: Forty nonleukoreduced PLT concentrates were stored at 20 to 24°C for up to 7 days. EBV genomes in B cells were quantified on Days 1 and 5. Viable white blood cells (WBCs) and T and B cells were quantified in 10 of 40 units on Days 1, 3, 5, and 7 of storage. For the Leukoreduction study, four pools of PLTs were leukoreduced within 24 hours of collection. B cells from before Leukoreduction and all peripheral blood mononuclear cells from after Leukoreduction were assayed for EBV. RESULTS: Viable WBCs and T cells were stable whereas viable B cells were reduced to 71% of the Day 1 level by Day 5. A total of 31 of 37 (83.8%) units were EBV positive. Although EBV genomes remained stable in most units, 12 of 37 units demonstrated a median of 5.1 (range 2- to 134)-fold increase in EBV genomes per 105 B cells on Day 5. For the Leukoreduction study, EBV genomes were detected in four of four pools before Leukoreduction with a median of 3.8 (range, 0.2-93.6) EBV genomes per 105 B cells. EBV genomes were not detected in any of the postLeukoreduction specimens. CONCLUSIONS: Seventy percent of B lymphocytes are viable on Day 5 of PLT storage. Although the mean number of EBV genomes remained stable, a subset of units had increased EBV genomes during storage. Leukoreduction removed polymerase chain reaction–detectable EBV genomes from PLT pools.
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efficacy of epstein barr virus removal by Leukoreduction of red blood cells
Transfusion, 2005Co-Authors: David W Rowe, Darrell J TriulziAbstract:BACKGROUND: Epstein-Barr virus (EBV) infection results in life-long carriage of latent virus in B lymphocytes in the majority of the adult population, including blood donors. The removal of EBV from red blood cell (RBC) components by Leukoreduction was assessed. STUDY DESIGN AND METHODS: Sixteen randomly selected fresh AS-5 units were leukoreduced by filtration. B lymphocytes from preLeukoreduction specimens and mononuclear cells (MNCs) from postLeukoreduction specimens were assayed for EBV DNA with sensitive real-time polymerase chain reaction (PCR). RESULTS: EBV genomes were detected in CD19+ B cells in 14 of 16 preleukoreduced RBC units. EBV genomic copy number in the units ranged from 0.18 to 96.84 per 105 B lymphocytes representing approximately 135 to 72,630 total EBV genomes per bag. Leukoreduction rendered all but one unit EBV-negative by PCR. The lone PCR-positive unit after Leukoreduction amplified 1.2 EBV genome copies from MNCs recovered from the entire unit of leukoreduced RBCs; this unit had the highest EBV viral load before Leukoreduction (72,630 EBV genomes). CONCLUSIONS: These results indicate that a 4-log reduction of EBV genomic copy number can be achieved with Leukoreduction of RBC units and renders most RBC units EBV-negative by sensitive PCR.