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Bacteria in platelet components (PC) may result in transfusion-related sepsis (TRS). Pathogen inactivation of PC with amotosalen (A-PC) can abrogate the risk of TRS and hence facilitate storage to 7 d. A randomized, controlled, double-blinded trial to evaluate the efficacy and safety of A-PC stored for 6-7 d was conducted. Patients were randomized to receive one transfusion of conventional PC (C-PC) or A-PC stored for 6-7 d. The primary endpoint was the 1 h corrected count increment (CCI) with an acceptable inferiority of 30%. Secondary endpoints included 1- and 24-h count increment (CI), 24-h CCI, time to next PC transfusion, red blood cell (RBC) use, bleeding and adverse events. 101 and 100 patients received A-PC or C-PC respectively. The ratio of 1-h CCI (A-PC:C-PC) was 0·87 (95% confidence interval: 0·73, 1·03) demonstrating non-inferiority (P = 0·007), with respective mean 1-h CCIs of 8163 and 9383; mean 1-h CI was not significantly different. Post-transfusion bleeding and RBC use were not significantly different (P = 0·44, P = 0·82 respectively). Median time to the next PC transfusion after study PC was not significantly different between groups: (2·2 vs. 2·3 d, P = 0·72). Storage of A-PCs for 6-7 d had no impact on platelet efficacy., (© 2011 Blackwell Publishing Ltd.)

Background: Whole blood and also buffy coats (BCs) can be held for a few hours or overnight before processing into blood components or platelet concentrates (PCs). Individual studies have reported a range of outcomes regarding in vitro variables for PCs prepared from fresh and stored whole blood. In this multicenter study, effects of storage of whole blood or BCs on the in vitro quality of PCs were studied., Study Design and Methods: The leukoreduced BC PCs were prepared from fresh BCs (2-8 hr after collection; fresh/fresh), from BCs at 20 to 24 hours after collection (fresh/stored), or from BCs prepared from whole blood stored for 20 to 24 hours (stored/fresh). PCs were stored on a flat-bed shaker at 20 to 24°C for 7 days. PCs were tested on Days 0 (only fresh/fresh), 1, 5, and 7 for in vitro quality. There were six participating centers that tested all three conditions with n = 6 per condition., Results: In comparison to fresh/stored and stored/fresh PCs, fresh/fresh PCs exhibited a lower platelet (PLT) count (Day 1-220 × 10(9) ± 70 × 10(9) vs. 324 × 10(9) ± 50 × 10(9) and 368 × 10(9) ± 56 × 10(9) PLTs/PC), lactate, pCO(2), and hypotonic shock response (HSR; Days 5 and 7; Day 7-50 ± 13% vs. 57 ± 12 and 63 ± 11%) and a higher pH, glucose, pO(2), and CD62P expression (than stored/fresh PCs only; Day 7-33 ± 10% vs. 28 ± 12 and 24 ± 11%; p < 0.05). No differences were observed for volume, swirling effect, white blood cell count, annexin V binding, or aggregation between these conditions., Conclusion: Based on PLT count, HSR, and PLT activation, PCs are best prepared after 20 to 24 hours hold of the whole blood or BCs., (© 2010 American Association of Blood Banks.)

Background and Objectives: The DiaMed Impact R tests platelet function under close to physiological flow conditions. The machine is designed to use whole blood but by adding back compatible red cells, it can be used to study stored platelet concentrates. To date, red cells ≤14 days old have been used. In this study, the effect on the assay of using red cells stored for up to 60 days was examined., Material and Methods: This study looked at buffy coat-derived platelet concentrates on day 2 of storage along with various stored red-blood-cells (RBC). To determine whether the age of the RBC is a factor in supporting adhesion and aggregation, platelets were assayed with either RBC stored between 2 and 60 days or with separated 'young' and 'old' red cell populations obtained using a centrifugation method and confirmed by percoll gradient analysis., Results: A statistically significant difference was observed between red-blood-cells stored for ≤20 days compared with those which have been stored for 21-60 days in respect of their ability to support platelet adhesion (SC) and aggregation (AS) (P<0·01). Separating red cells by centrifugation into top (young population) and bottom (old population) showed that the effect of storage was much greater than was any difference between young and old at the individual time-points e.g. 'young' red cells from stored units were poorer at supporting platelet adhesion and aggregation than 'young' red cells from fresh units., Conclusion:   Results suggest that the red cells should be stored for less than 21 days when using this assay. This assay may also allow assessment of red cell functionality., (© 2010 The Author(s). Vox Sanguinis © 2010 International Society of Blood Transfusion.)

Summary: New platelet storage systems, such as changes in the plastic of the storage bags, require validation. In this study, pooled buffy coat platelets stored in Fresenius/NPBI polyolefin bags were compared with those stored in Fresenius/NPBI butyryl-trihexyl citrate (BTHC) plasticized polyvinyl chloride (PVC). The CompoSelect thrombocyte polishing filter system (1000 mL polyolefin bag) and the CompoStop F730 system (1300 mL BTHC-PVC bag) were used to prepare paired, plasma-suspended, buffy coat platelet concentrates. Samples were taken up to day 7 for in vitro analysis. In a separate experiment, 12 units were prepared using the CompoStop F730 system and samples taken after leucofiltration for FXIIa assay. By day 7, platelet concentrates stored in BTHC-PVC demonstrated significantly higher pH levels (7.32 +/- 0.05 vs. 7.26 +/- 0.05) and a greater degree of cell lysis as shown by increased lactate dehydrogenase levels (497 +/- 107 vs. 392 +/- 81 U L(-1)). The supernatants contained higher concentrations of soluble P-selectin and the chemokine 'regulated on activation, normal T-cell expressed and presumably secreted', which are released from the alpha-granules during activation. The ATP concentrations were significantly lower in BTHC-PVC. Platelet counts, mean platelet volume and hypotonic shock response were similar for both bags. FXIIa antigen concentrations were 0.6 +/- 0.2 ng mL(-1) indicating that activation of the contact factor pathway had not occurred. Although the CompoStop F730 leucoreduction filter did not activate the contact system, platelets stored in 100% plasma in BTHC-PVC bags demonstrated different in vitro characteristics from those stored in polyolefin. Further work is required to demonstrate whether these differences will affect in vivo recovery and survival.

Background: This study was designed to determine which in vitro assays would be most useful for studying the effects of cold storage on platelet concentrates and to establish an in vivo model for platelet recovery and survival., Study Design and Methods: Paired, plasma-suspended, leucoreduced, buffy-coat-derived platelet concentrates were stored either at 22 or 4 degrees C. Prior to storage and after 18 h, 5 days and 7 days, samples were taken and various assays were performed. On day 6, in vivo studies were carried out using a model system. Galactosylation of the platelets, prior to cold storage, was also tested., Results: Hypotonic shock response, collagen-induced aggregation, RANTES and P-selectin binding site measurements demonstrated differences between platelets stored at 22 and 4 degrees C. The glycocalicin assay was able to demonstrate microvesicle formation at 4 degrees C. The in vivo model showed that there was at least a 50% decrease in recovery and survival when the platelets were stored in the cold. Galactosylation did not improve these results., Conclusions: Several assays, both in vitro and in vivo, were able to detect differences in platelet-storage characteristics and in vivo recovery and survival in a model system. Galactosylation did not correct these cold-induced changes.

The mechanisms involved in storage-induced damage in platelets are not well understood, but membrane signalling via Ca2+ ion flux may affect mitochondrial H+ gradients and metabolism and the intrinsic pathways of cell death, platelet survival and function. In this study, the effects of blood bank storage conditions, including reduced plasma concentration and interrupted agitation, were evaluated in platelets from 136 healthy donors. Mitochondrial membrane potential (DeltaPsim), an indicator of intrinsic cell death, and its sensitivity to Ca2+ ionophore A23187, were monitored using JC-1 by flow cytometry and fluorescence microscopy. Platelet survival was examined using lactate dehydrogenase release, annexin V binding and caspase-3/7 activity. Decreased plasma concentration and interrupted agitation affected DeltaPsim and caspase-3/7. Over 7 days in 30% plasma DeltaPsim showed a significant reduction (86.3 +/- 1.1% platelets with polarised mitochondria day 1; 79.9 +/- 2.1% day 5; 75.1 +/- 3.8% day 7, P = 0.01 day 1 vs. day 7). Whilst DeltaPsim in agitated platelets in 100% plasma was unchanged up to day 7, interruption of agitation was associated with a 44% reduction in the proportion of platelets with polarised mitochondria after 5 days (56 +/- 11%). The Ca2+ sensitivity of DeltaPsim changed earlier: 5 microM A23187 caused a 20-30% change in the fraction of platelets with polarised mitochondria by day 5. Ca2+ sensitivity also increased during interrupted agitation and reduced plasma concentration. DeltaPsim also correlated with indicators of platelet death, caspase-3 activity and annexin V binding (correlation coefficients of 0.8). In conclusion, changes in Ca2+-sensitive DeltaPsim are involved in the initiation of storage-induced cell death signals that influence platelet count and function in vivo.

Background and Objectives: The aim of this study was to investigate the effects of extended storage of pooled random platelets in SSP+ additive solution (MacoPharma)., Materials and Methods: Eight buffy coat-derived, pooled, leucoreduced platelet concentrates were prepared in 75% SSP+, 25% plasma using Fresenius/NPBI Composelect thrombocyte polishing filter (TPF) systems. Platelet concentrates were stored for 19 days in polyolefin storage bags and samples for in vitro analysis were taken at various time-points during storage., Results: Platelet yields were lower than seen routinely when platelets are prepared in 100% plasma. The in vitro quality of the platelets stored in SSP+ was maintained until day 9. Glucose was depleted by day 12 and this was accompanied by a rapid fall in pCO2, a rise in pO2 and a cessation of lactate production. ATP and bicarbonate concentrations fell, the platelets began to swell and the ability to swirl decreased. Soluble P-selectin, glycocalicin, and regulated on activation, normal, T-cell expressed, and secreted (RANTES) concentrations increased, as did P-selectin expression. Loss of platelets and an increase in lacate hydrogenase concentration indicated that lysis had occurred. However, the pH remained between 6.4 and 7.4., Conclusions: The results suggest that SSP+ could be used for platelet storage for up to 9 days. However, the preparation of platelets in the additive requires some optimization. In vivo studies are required to confirm these in vitro results.

Background and Objectives: Extent of shape change (ESC) and hypotonic shock response (HSR) have been widely used to characterize the in vitro function of platelets and have been shown to correlate with in vivo viability. These assays have been routinely performed using platelet-poor plasma (PPP) as the test sample diluent. Because of the increasing popularity of storing platelets in synthetic media, it is important to understand the effects of using these synthetic media as test diluents for ESC and HSR measurements. The objective of this study was to determine the effect of using platelet storage solutions vs. plasma for the in vitro testing of ESC and HSR., Materials and Methods: Six laboratories participated in this study. Platelets were prepared by apheresis, the platelet-rich plasma (PRP) method, or derived from buffy-coats. Each platelet preparation was divided, half being stored in plasma and the other half in storage solution. ESC and HSR testing were performed in duplicate on days 1 and 5, using each of three diluents: autologous plasma; fresh-frozen plasma; or storage solution., Results: For both ESC and HSR, dilutions made in each of the three diluents yielded significantly different results. Dilutions made in storage solutions were more than 30% lower for ESC and HSR than those made in autologous plasma (P < 0.0001). Dilutions made in thawed fresh-frozen plasma were more than 16% lower for ESC and HSR than those made in liquid autologous plasma (P < 0.0005)., Conclusions: ESC and HSR test results are significantly affected by the test diluent. Platelets should be diluted in plasma (preferably autologous) for the in vitro testing of ESC and HSR, regardless of the media in which they are stored.

Background and Objectives: In a preliminary study, the presence of potassium and magnesium in a modified synthetic medium (PAS-III) was found to have a significant influence on platelet metabolism (using apheresis-derived, as well as buffy-coat-derived platelets) when compared with standard PAS-III. The differences included reduced glycolysis, as evidenced by lower consumption of glucose and lower production of lactate, but also better preservation of pH and hypotonic shock response reactivity. The results suggested that storage in modified PAS-III containing 20% plasma was comparable to storage in standard PAS-III containing 30% plasma. To confirm the preliminary results and to evaluate the effects of different preparation protocols, an international multicentre study, which included 11 different sites, was conducted., Materials and Methods: Platelets from 30 pools of approximately 20 buffy coat (BC) units each and 24 pooled apheresis platelet units were aliquoted for storage in plasma (reference) or synthetic medium using either a specific additive solution (PAS-III) containing 30% plasma or a modification of PAS-III containing 5.0 mm potassium and 1.5 mm magnesium (PAS-IIIM) and either 30% or 20% plasma. Units were stored at room temperature with agitation for 7 days during which in vitro testing was carried out for biochemical, haematological and functional parameters., Results: Storage of platelets in PAS-IIIM resulted in a reduction in the rate of glycolysis and better retention of pH and hypotonic shock response reactivity. Storage in PAS-IIIM containing 20% plasma appeared to result in the retention of in vitro properties, similar to those observed during storage in standard PAS-III containing 30% plasma., Conclusions: The results of this study confirm the preliminary results. Similar results were seen with platelets prepared by BC and apheresis methods, despite differences in equipment, the preparation technique and in the final platelet contents achieved in the platelet units. Storage of platelets in PAS-IIIM should be considered to improve platelet function and allow plasma reduction to 20%., (Copyright 2003 Blackwell Publishing)

Background: Recent studies using a time-resolved fluoroimmunoassay method (dissociation-enhanced lanthanide fluoroimmunoassay) showed that platelets and plasma are the main reservoir of the normal isoform of cell-associated prion protein (PrPc) in human blood. The aims of the present study were to monitor PrPc levels in various fractions of apheresis platelets during storage by using the DELFIA method and to assess the association of this release with alpha-granule protein ss-thrombo-globulin and cytoplasmic LDH., Study Design and Methods: Units of apheresis platelets (n = 6) were obtained from volunteer donors by the use of a cell separator and stored up to 10 days. Samples (7-9 mL) were aseptically collected from each unit on storage Days 1, 2, 3, 4, 5, 8, and 10. Platelet-poor plasma and apheresis platelets were prepared and the former split into two fractions, one centrifuged at 40,000 x g for 2 hours at 4 degrees C to remove microparticles. The spun microparticles, apheresis platelets and platelet samples, platelet-poor plasma, and high-spun plasma fractions were stored in a frozen state until they were tested., Results: The results showed that the mean overall levels of PrPc throughout storage remained within 15 percent of Day 1 levels. In contrast, the mean cellular levels in platelets significantly decreased to 46 percent of Day 1 levels by Day 10 of storage (p<0.01), while the corresponding levels in plasma significantly rose as much as 329 percent (p<0.01). Moreover, although microparticle-bound PrPc was released during storage, it was increasingly superseded by soluble protein. PrPc and ss-thrombo-globulin release exhibited very similar patterns (p<0.01). In contrast, LDH showed a significant increase in high-spun plasma only toward the end of the storage period (p<0.01)., Conclusion: These results indicate that PrPc is released from platelets during the storage of apheresis platelets and that this release is probably due mainly to platelet activation and alpha-granule release in the first few days of storage. Moreover, the released PrPc is increasingly composed of soluble proteins, as the storage period exceeds 5 days.

A range of membrane-active drugs were tested for their ability to prevent beta-thromboglobulin and platelet factor 4 release from freshly collected blood platelets. While all the drugs tested could inhibit collagen-induced platelet aggregation, only a few, notably procaine and the anti-malarial drugs chloroquine, hydroxychloroquine, camoquine and quinacrine (mepacrine), effectively prevented the alpha-granule release reaction.

Five monoclonal antibodies to human von Willebrand factor were selected for characterization from 18 produced in murine hybridomas. All showed a high and specific affinity for human von Willebrand factor (vWf) but exhibited little if any cross-reaction with sera from other species. The antibodies defined four epitopes on vWf, none of which were involved in platelet binding. Binding of two distinct antibodies at one of these epitopes was associated with enhancement of the rate of vWf-dependent platelet agglutination in the presence of ristocetin. This effect was more noticeable when cryosupernatant plasma was used in place of normal plasma as the source of vWf, and was not explicable simply in terms of antibody-induced cross-linking of vWf.

The possible correlation between soluble immune factors and platelet and coagulation factors has been evaluated in Type 1 diabetic patients with and without proliferative retinopathy, and in non-diabetic controls. Soluble immune complexes, platelet factor IV (PF4), beta-thromboglobulin, fibrinogen, factor VIII related antigen and anti-thrombin III were significantly increased in Type 1 diabetic patients with retinopathy as compared to non-diabetic controls. Fibrinogen and anti-thrombin III were also higher in those patients with retinopathy compared to those without retinopathy. A significant correlation was found between positive values of soluble immune complexes and increased levels of PF4 and beta-thromboglobulin in diabetic patients with retinopathy. The presence of soluble immune complexes and insulin-anti-insulin complexes was associated with a significantly greater number of elevated haemostatic factors in retinopathic patients. Our findings suggest that the interaction of platelets and soluble immune complexes or insulin-anti-insulin complexes may be pathologically relevant to the development of diabetic retinopathy.

Three human volunteers were injected with a range of doses of pentosan polysulphate, SP54, i.v. or s.c. A competitive binding assay (CBA) for sulphated polysaccharides was used to detect circulating SP54 after doses as low as 1 mg i.v. and a linear relationship was observed between the peak plasma concentration of SP54 measured by CBA and the administered dose. A comparison was made between the clearance of SP54 measured by CBA and its anticoagulant and lipolytic activities. SP54 was detectable by CBA after doses which caused no alteration in activated partial thromboplastin time (APTT) or anti-factor Xa activity but after which a small increase of lipase activity was measurable. After SP54 at 10 mg i.v. or 100 mg s.c. anti-factor Xa activity was 4-6 times greater than would be expected from the in vitro activity of the concentrations of SP54 measured by CBA. Like heparin and other heparin analogues, SP54 caused an increase in plasma concentrations of platelet factor 4 (PF4) without a concomitant rise in beta-thromboglobulin (beta-TG). It is concluded that the newly developed CBA will provide a more sensitive means than conventional bioassays for the determination of plasma concentrations of SP54.

Background and Objectives Pathogen inactivation (PI)-treated plasma and platelets are increasingly becoming the products of choice, where licensed. This review summarizes the clinical evidence available for licensed component PI technologies and red cell PI under development. Materials and Methods Available literature on licensed technologies was reviewed. Results For the plasma and platelets technologies available, evidence for the inactivation of most pathogens is good, except for certain nonenveloped viruses. Clinical trials and haemovigilance programmes suggest the observed loss of potency is of little clinical significance, with some technology-specific exceptions. Concerns over adverse toxicological effects or neoantigen formation have not been confirmed for currently licensed products. Conclusion While platelet PI has been adopted to reduce bacterial contamination, the ability of PI methods to replace testing for emerging bloodborne infections, or as a substitute for selective pathogen testing, gamma-irradiation or even leucodepletion, make adoption of PI for components increasingly attractive. [ABSTRACT FROM AUTHOR]

Background and Objectives The aim of this study was to investigate the effects of extended storage of pooled random platelets in SSP+ additive solution (MacoPharma). Materials and Methods Eight buffy coat-derived, pooled, leucoreduced platelet concentrates were prepared in 75% SSP+, 25% plasma using Fresenius/NPBI Composelect thrombocyte polishing filter (TPF) systems. Platelet concentrates were stored for 19 days in polyolefin storage bags and samples for in vitro analysis were taken at various time-points during storage. Results Platelet yields were lower than seen routinely when platelets are prepared in 100% plasma. The in vitro quality of the platelets stored in SSP+ was maintained until day 9. Glucose was depleted by day 12 and this was accompanied by a rapid fall in pCO2, a rise in pO2 and a cessation of lactate production. ATP and bicarbonate concentrations fell, the platelets began to swell and the ability to swirl decreased. Soluble P-selectin, glycocalicin, and regulated on activation, normal, T-cell expressed, and secreted (RANTES) concentrations increased, as did P-selectin expression. Loss of platelets and an increase in lacate hydrogenase concentration indicated that lysis had occurred. However, the pH remained between 6·4 and 7·4. Conclusions The results suggest that SSP+ could be used for platelet storage for up to 9 days. However, the preparation of platelets in the additive requires some optimization. In vivo studies are required to confirm these in vitro results. [ABSTRACT FROM AUTHOR]

The aim of this study was to assess the separation of whole blood into red cells and plasma by using the Sangofer® device, which is a gravity-fed, hollow-fibre system. The components would then be compared with those produced by the use of more elaborate technical equipment. Ten whole-blood units were leucoreduced by using a WBF2 filter and immediately separated into red cells and plasma by using the Sangofer® blood-separation device. Red cells were stored in additive solution and tested on days 1 and 42. The plasma was assayed for levels of various coagulation factors and for markers of both coagulation and complement activation. The red-cell parameters were similar to those obtained when routine centrifugation methods were used. The filter did not cause haemolysis. Levels of plasma factor VIII and factor XI were lower than those seen in routinely produced leucoreduced plasma units but there was no evidence of activation of the coagulation and complement systems. The Sangofer® device is simple and straightforward to use and eliminates the need for both centrifugation and automated separation steps during the processing of whole blood into red cells and plasma components. Minor changes are required to make the procedure easier to incorporate into routine use. [ABSTRACT FROM AUTHOR]

When platelet concentrates (PCs) are shipped from one centre to another, they may remain unagitated for a considerable period of time. It was therefore our aim to study the effects of interruption of agitation on thein vitroparameters of PCs stored in platelet additive solutions.In this multicentre study, PCs were prepared either by apheresis or from pooled buffy coats, paired to minimize donor-dependent differences, and aliquoted into 3 units with a‘low concentration’ (≈ 1 × 109 platelets/ml; groups A, B and C) and 3 units with a‘high concentration’ (≈ 2 × 109 platelets/ml; groups D, E and F). The final composition of the storage medium was 30% plasma and 70% additive solution in all PCs. Either PASIIIM or Composol was used as the additive solution. Agitation was interrupted for 2 days (between days 3 and 5, groups A and D), or for 4 days (between days 1 and 5, groups B and E), and continuous agitation served as the reference (groups C and F). A number ofin vitroparameters were used for testing on days 1, 5 and 7.On day 7, reference units C and F in PASIIIM had significantly higher pH values than the study units in PASIIIM, but all retained a pH of> 6·5 at 37 °C. Hypotonic shock response (HSR) results were significantly lower in the high concentration/4-day interruption group (E) than in the other groups. The low-concentration groups in PASIIIM, with agitation interrupted for either 2 days (group A) or 4 days (group B), did not have HSR values significantly different from the respective references. Study groups A, B, D and E in Composol, a solution lacking phosphate, had a pH of≈ 6·5 on day 7, which was significantly lower than that of the references and of the corresponding units in PASIIIM. The pH values were>7·0 in reference groups C and F in Composol, not significantly different from those in PASIIIM. HSR values were also significantly lower in the Composol study groups. On the other hand, the reference Composol groups showed results similar to units in PASIIIM.PCs in PASIIIM additive solution with a platelet concentration of≈ 1 × 109/ml can sustain 4 days without agitation. Phosphate may be of importance in maintaining goodin vitrocharacteristics during interruption of agitation. [ABSTRACT FROM AUTHOR]