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Background: Pathogen reduction (PR) of platelet concentrates (PCs) contributes to the safety of platelet (PLT) transfusion by reducing the risk of transfusion-transmitted infections and transfusion-associated graft-versus-host disease. In vitro quality of pathogen-reduced double-dose PC (PR-PC) made of eight whole blood (WB)-derived buffy coats (BCs) were evaluated., Methods: Eight small-volume WB BCs from donors with at least 200 × 10 9 PLT/L were pooled with an additive solution to produce double-dose PCs (DD-PCs), which were treated with amotosalen/ultraviolet A light in a dual storage processing set, yielding 2 units of PR-PC. Quality controls were undertaken as per European Directive for the Quality of Medicines (EDQM) guidelines. PLT recovery rates were measured. Production costs and savings were compared over the 3 years before and after PR implementation., Results: In the pre-PR period, 19 666 PCs were produced, compared to 17 307 PCs in the PR period. Single BC in the PR period had 41 ± 2 mL, hematocrit 0.39 ± 0.04 and 1.06 ± 0.18 × 10 11 PLTs, and showed a recovery of 91% ± 8%. After pooling, separation, PR treatment of DD-PC, and splitting, each single PC had 189 ± 6 mL with 2.52 ± 0.34 × 10 11 PLTs, compared to 2.48 ± 0.40 in the pre-PR period. The PLT recovery rate after PR was 87% ± 14%. EDQM requirements were met. An increase of about €12 (+7.5%) per PC from the pre-PR to the PR period was identified., Conclusion: A new production method resulting in two PR-PCs made from pools of 8 BCs with use of one PR set was successfully introduced, and our experience of nearly 3 years demonstrated the high efficacy and in vitro quality of the PR-PCs obtained., (© 2020 The Authors. Transfusion published by Wiley Periodicals LLC. on behalf of AABB.)

Background: UVC illumination of agitated platelet concentrates (PCs) inactivates pathogens and white blood cells by modifications of their nucleic acids. Related effects on mitochondrial DNA (mtDNA) in platelets serve as a basis for an efficient monitoring suited for routine quality control (QC) of this purely physical pathogen reduction technology., Study Design and Methods: Samples from PCs (n = 530) were tested with an established LightCycler PCR (LC PCR) for QC of the UVC procedure. RNR2 and TRNK/ATP8 genes were sequenced in the PCs (n = 21) with out-of-specification results in the LC PCR. A digital droplet PCR (ddPCR) was developed to minimize the outliers and cross-validated by testing the 530 PCs. The ddPCR was further evaluated in a subgroup of 300 PCs without mtDNA extraction and in samples from systematic variations of UVC dose and agitation speed., Results: Apheresis PCs (n = 380) resulted in 5.3% outliers in LC PCR versus only 0.7% in buffy coat pool PCs (n = 150). Sequencing of these outliers revealed single-nucleotide polymorphisms in the primer- and probe-binding sites of LC PCR. The development of a ddPCR assay with modified probe sequences reduced the outliers to 0.4%. The ddPCR analysis of PCs both with and without mtDNA extraction demonstrated low intra- and interassay variabilities and congruent results also compared to LC PCR. Experiments varying the UVC dose and the agitation speed demonstrated that the ddPCR results closely reflect functional effects of the UVC treatment., Conclusion: The ddPCR assay offers a valid and reliable tool for QC of routine production of the UVC-treated PCs as well as for monitoring treatment conditions during optimization of the UVC procedure., (© 2020 AABB.)

Background: Three methods of leukoreduction (LR) are used worldwide: filtration, buffy coat removal (BCR), and a combination of the previous two methods. Additionally, there are a number of additive solutions (ASs) used to preserve red blood cell (RBC) function throughout storage. During RBC storage, proinflammatory activity accumulates; thus, we hypothesize that both the method of LR and the AS affect the accumulation of proinflammatory activity., Study Design and Methods: Ten units of whole blood were drawn from healthy donors, the RBC units were isolated, divided in half by weight, and leukoreduced by: 1) BCR, 2) filtration, or 3) BCR and filtration (combination-LR); stored in bags containing AS-3 per AABB criteria; and sampled weekly. The supernatants were isolated and frozen (-80°C). RBC units drawn from healthy donors into AS-1-, AS-3-, or AS-5-containing bags were also stored and sampled weekly, and the supernatants were isolated and frozen. The supernatants were assayed for neutrophil (PMN)-priming activity and underwent proteomic analyses., Results: Filtration and combination LR decreased priming activity accumulation versus buffy coat LR, although the accumulation of priming activity was not different during storage. Combination LR increased hemolysis versus filtration via proteomic analysis. Priming activity from AS-3 units was significant later in storage versus AS-1- or AS-5-stored units., Conclusions: Although both filtration and combination LR decrease the accumulation of proinflammatory activity versus buffy coat LR, combination LR is not more advantageous over filtration, has increased costs, and may cause increased hemolysis. In addition, AS-3 decreases the early accumulation of PMN-priming activity during storage versus AS-1 or AS-5., (© 2018 AABB.)

Background: Bacterial contamination of platelet concentrates (PCs) poses the major posttransfusion infectious risk in developed countries. The aerobic microorganism most frequently isolated from PCs is coagulase-negative Staphylococcus epidermidis, a normal inhabitant of the human skin, which has been involved in fatal transfusion reactions worldwide., Case Report: In September 2014, a splenectomized elderly male patient, suffering from leukemia, was transfused with two 5-day-old buffy coat platelet (PLT) pools. The patient returned to emergency on the same day with a low-grade fever. He was bacteremic and died on the next day. Microbiology and molecular testing of a blood sample from the patient and one of the PCs yielded the same S. epidermidis strain. Further analysis demonstrated that this S. epidermidis isolate displays a biofilm-positive phenotype in PCs., Discussion: At Canadian Blood Services, PCs are screened for bacterial contamination with the BacT/ALERT system (bioMérieux) at approximately 24 hours postcollection. The implicated PC had been tested and yielded a false-negative culture result. A titration experiment indicated that, at the time of screening, the contaminated PC had a titer of less than 0.74 colony-forming units (CFU)/mL (<227 CFUs/unit) of S. epidermidis. Mathematical models have predicted that up to 70% of PCs contaminated with coagulase-negative staphylococci at concentrations of 0.02 CFU/mL can be missed by BacT/ALERT screening., Conclusion: Despite several mitigation strategies, false-negative cultures with current PLT screening practices still occur. This report creates awareness of the pathogenicity of opportunistic S. epidermidis, a low-virulence organism, in susceptible patients who may not develop a typical transfusion reaction., (© 2015 AABB.)

Background: Platelet additive solutions (PASs) facilitate plasma recovery and may reduce the risk of plasma-associated adverse transfusion reactions. Whereas current apheresis techniques are able to produce platelet concentrates (PCs) with reduced residual plasma volumes, it is still technically challenging to prepare PCs with plasma levels less than 20% from whole blood. This study aimed to evaluate the feasibility of producing buffy coat (BC)-derived platelets (PLTs) with as little as 10% residual plasma and to test the ability of PASs to preserve PLT quality under these conditions., Study Design and Methods: A pool-and-split design was used to evaluate the in vitro quality of semiautomatically prepared BC-derived PCs stored for 7 days in either SSP+ (PAS-IIIM) or Intersol-G (glucose-containing PAS) with 10% to 30% residual plasma in three phases: 1) 30% plasma (SSP+, Intersol-G), 2) 20 and 15% plasma (SSP+, Intersol-G), and 3) 10% plasma (Intersol-G)., Results: The different plasma-reduced PAS-PC types were comparable in volume, PLT concentration, and PLT yield and met all regulatory requirements. The in vitro quality variables of PLTs suspended in 20% or less residual plasma were comparable to those of PLTs stored in 30% plasma for both PAS types. PLT activation was slightly higher with SSP+ than with Intersol-G. The quality of PCs suspended in Intersol-G with 10% plasma was well maintained during storage., Conclusions: Preparation of BC-derived PCs with minimal plasma concentrations as low as 10% is feasible. Late-generation PASs satisfactorily preserve the in vitro quality of such PCs during storage., (© 2015 AABB.)

Background: Platelet concentrates (PCs) can be prepared using three methods: platelet (PLT)-rich plasma, apheresis, and buffy coat. The aim of this study was to obtain a comprehensive data set that describes metabolism of buffy coat-derived PLTs during storage and to compare it with a previously published parallel data set obtained for apheresis-derived PLTs., Study Design and Methods: During storage we measured more than 150 variables in 8 PLT units, prepared by the buffy coat method. Samples were collected at seven different time points resulting in a data set containing more than 8000 measurements. This data set was obtained by combining a series of standard quality control assays to monitor the quality of stored PLTs and a deep coverage metabolomics study using liquid chromatography coupled with mass spectrometry., Results: Stored PLTs showed a distinct metabolic transition occurring 4 days after their collection. The transition was evident in PLT produced by both production methods. Apheresis-derived PLTs showed a clearer phenotype of PLT activation during early days of storage. The activated phenotype of apheresis PLTs was accompanied by a higher metabolic activity, especially related to glycolysis and the tricarboxylic acid cycle. Moreover, the extent of the activation differed between bags resulting in interbag variability in the storage lesion of apheresis-prepared PLTs. This may be related to donor-related polymorphism., Conclusion: This study demonstrated two discrete metabolic phenotypes in stored PLTs prepared with both apheresis and buffy coat methods. PLT activation occurs during the first metabolic phenotype and might lead to a low reproducibility of the apheresis PCs., (© 2014 AABB.)

Background: Extracorporeal photopheresis (ECP) is commonly used to treat patients with graft-versus-host disease (GVHD) and lung transplant rejection (LTR) in our institution. The quantitative relationship between the number of white blood cells treated during ECP and the cell count in peripheral blood is unclear., Study Design and Methods: Patients with GVHD and LTR receiving ECP with either UVAR XTS or CELLEX (Therakos) were prospectively recruited for this study. A complete cell count with differential was performed on preprocedural peripheral blood and samples from the collected buffy coats. Correlation analysis and linear regression were performed between cell counts in peripheral blood and buffy coat. Collection efficiency was compared between UVAR XTS and CELLEX., Results: In all 52 patients, lymphocyte counts in buffy coat and peripheral blood showed strong correlation (r values were 0.85 and 0.983 for UVAR XTS and CELLEX, respectively; p < 0.001) with slopes of 2 and 5.1 for UVAR XTS and CELLEX, respectively (p < 0.001). The quantitative relationship remained robust in patients stratified by diagnoses. Monocytes also showed consistent correlation and linearity, but not neutrophils or combined white blood cells, red blood cells, or platelets. CELLEX enriched approximately twice as many lymphocytes and monocytes than UVAR XTS per procedure (p < 0.001)., Conclusion: The preprocedural peripheral lymphocyte count can predict the number of lymphocytes within the buffy coat collected during ECP, which may justify the use of peripheral lymphocyte count as a surrogate for the cell dose treated per procedure. Peripheral monocyte counts may serve as an alternative. CELLEX is more efficient in collecting lymphocytes and monocytes than UVAR XTS under conditions tested., (© 2013 American Association of Blood Banks.)

Background: We evaluated the effect of treating platelets (PLTs) using ultraviolet (UV)C light without the addition of any photosensitizing chemicals on PLT function in vitro and PLT recovery and survival in an autologous radiolabeled volunteer study., Study Design and Methods: For in vitro studies, pooled or single buffy coat-derived PLT concentrates (PCs) were pooled and split to obtain identical PCs that were either treated with UVC or untreated (n = 6 each) and stored for 7 days. PLT recovery and survival were determined in a two-arm parallel autologous study in healthy volunteers performed according to BEST guidelines. UVC-treated or untreated PCs (n = 6 each) were stored for 5 days and were compared to fresh PLTs from the same donor., Results: There were no significant differences on Day 7 of storage between paired UVC-treated and control PC units for pH, adenosine triphosphate, lactate dehydrogenase, CD62P, CD63, PLT microparticles, and JC-1 binding, but annexin V binding, lactate accumulation, and expression of CD41/61 were significantly higher in treated units (p < 0.05). Compared with control units, the recovery and survival of UVC-treated PC were reduced after 5 days of storage (p < 0.05) and when expressed as a percentage of fresh values, survival was reduced by 20% (p = 0.005) and recovery by 17% (p = 0.088)., Conclusion: UVC-treated PLTs stored for 5 days showed marginal changes in PLT metabolism and activation in vitro and were associated with a degree of reduction in recovery and survival similar to other pathogen inactivation systems that are licensed and in use., (© 2012 American Association of Blood Banks.)

Background: Blood operators have taken measures to reduce transfusion-related acute lung injury (TRALI). We classified suspected TRALI cases reported to Canadian Blood Services from 2001 to 2009 and assessed the impact of TRALI reduction measures., Study Design and Methods: Using Canadian Consensus Conference definitions, cases were reviewed by two experts or, from 2006 to 2009, a TRALI Medical Review Group (TMRG). Detection of HLA antibodies was performed using the Luminex system starting in 2008. Measures implemented from 2007 to 2009 included use of predominantly male plasma, suspension of buffy coat platelets in male plasma, and deferral of females with a pregnancy history from plateletpheresis. The buffy coat production method was implemented from 2005 to 2008., Results: Reporting of all suspected TRALI cases, as well as cases classified as definite or possible, increased from 2001 to 2004, was stable from 2004 to 2007, and declined in 2008 to 2009. The decline was most marked for plasma-associated cases, but occurred for all components. TMRG consensus on classification was achieved in 56% of cases. Cases identified as definitive or possible TRALI were significantly more likely to have donor antibody against a corresponding recipient antigen, compared to other cases., Conclusion: Hemovigilance data demonstrated an initial increase in TRALI cases, likely due to increased adverse event reporting and awareness of TRALI, followed by a decrease in cases related to all components. TRALI prevention measures and possibly the switch to the buffy coat production method may have contributed to the decline. Classification of cases remains challenging., (© 2011 American Association of Blood Banks.)

Background: Besides white blood cell antibodies in plasma-rich products, another cause of transfusion-related acute lung injury (TRALI) could be release of biologically active substances during storage of cellular blood products. We aimed to investigate the association of storage time and risk of TRALI for different product types., Study Design and Methods: We compared storage time of blood products transfused within 6 hours before the onset of TRALI to storage time of a representative sample of all blood products transfused in the Netherlands. Generalized linear models were used to correct for confounding variables., Results: Platelets (PLTs) in plasma transfused to TRALI patients were stored for 0.7 (95% confidence interval [CI], 0.073 to 1.3) days longer than those transfused to controls. The relative risk of TRALI, after receiving PLTs stored for 4 or 5 days, compared to 3 days or less, was 5.8 (95% CI, 0.99 to 110) and increased to 6.3 (95% CI, 1.1 to 118) after more than 5 days (i.e., 6 or 7 days)., Conclusions: While longer storage of buffy coat-derived PLTs was associated with an increased risk of TRALI, storage of plasma for up to 2 years and red blood cells for up to 35 days was not associated with the risk of TRALI., (© 2011 American Association of Blood Banks.)

Background: With buffy coat (BC) processing of whole blood (WB) donations, increase in WB storage time to facilitate overnight holding before the separation of blood components would be a logistically attractive development. This study undertakes a comparative in vitro characterization of blood components prepared from WB samples that were either processed within 8 hours or stored overnight at room temperature before processing by the BC method., Study Design and Methods: The WB units (400 mL) collected were either processed within 8 hours (fresh blood) or stored overnight (overnight blood) at room temperature. WB units were separated into individual-component red blood cells (RBCs), BC, and plasma. The in vitro quality of these blood components (RBCs, pooled platelet concentrates [PCs], and plasma) was analyzed during storage., Results: Levels of 2,3-diphosphoglycerate (2,3-DPG) were found to be significantly lower immediately after processing, compared with the fresh WB samples, in RBCs that had been separated from an overnight-hold sample. However, this difference was not apparent after 14 days of storage. In pooled PCs, measurements for glucose, lactate, PO(2), PCO(2), extent of shape change, and hypotonic shock response were similar. The platelet yield in PCs prepared from an overnight-hold WB sample was significantly higher, while CD62P expression and annexin V binding were lower (p < 0.05). For frozen plasma (FP), no significant differences were observed for the coagulation factors (F)II, FVII, FV, F IX, FX, and FXI; fibrinogen; and von Willebrand factor content between the 8- and 24-hour FP. The FVIII was the component that was most sensitive to the prolongation of production time and it only had 80% of the activity of the 8-hour FP., Conclusion: These data suggest that blood components (RBCs, pooled PCs, and FP) separated from WB that has been stored overnight at room temperature by the BC method are of acceptable quality., (© 2011 American Association of Blood Banks.)