57. Jahrestagung der Deutschen Gesellschaft für Transfusionsmedizin und Immunhämatologie

Dr. med.

Vorsitz

Prof. Dr. med.

Platelet toll-like receptor 2-dependent immune function is preserved under short-term refrigeration of platelets

Blood Components

Die thrombozytäre Toll-like-Rezeptor 2-vermittelte Immunfunktion bleibt unter kurzzeitiger Kühl-Lagerung von Thrombozyten erhalten

Recently, it was demonstrated that short-term refrigeration of platelets for 1 h or 2 h - either in platelet-rich-plasma (PRP) or in stored platelet concentrates (PC) - is sufficient to generate platelets with higher hemostatic responsiveness. However, refrigeration may have impact on non-hemostatic functions of platelets. Therefore, the intention of this study was to investigate the effect of short-term refrigeration on specific toll-like receptor 2-mediated platelet functions, as previously explored.PRP, freshly prepared from healthy donors, was stored at cold temperature (CT, 2-6 °C) for 1 or 2 h, or alternatively, at room temperature (RT, 20-24 °C), followed by analysis of TLR2-dependent platelet integrity. TLR2 expression, intracellular reactive oxygen species (ROS) production in H2DCFDA-loaded platelets and uptake of fluorescence-labeled synthetic TLR2 ligand Pam3CSK4, and fluorophore-conjugated fibrinogen were evaluated by flow cytometry. TLR2-induced release of chemokines PF4 (platelet factor 4), NAP-2 (neutrophil-activating peptide-2), sCD40L (soluble CD40 ligand), RANTES (Regulated on activation, normal T cell expressed and secreted) and PDGF (platelet-derived growth factor) was measured by immunoassays.Compared to RT, TLR2 expression shows a slightly increasing tendency under storage at CT. The uptake of fibrinogen and the uptake of ligand Pam3CSK4 in unstimulated and Pam3CSK4-stimulated platelets is comparable at RT and CT. The basal and Pam3CSK4-induced production of intracellular ROS remains unchanged at both temperatures. Accordingly, the release of chemokines PF4, NAP-2, sCD40L, RANTES and PDGF is not relevantly affected by storage at CT.Short-term refrigeration is an opportunity to generate platelets with higher hemostatic reactivity without tampering the function of platelet TLR2. Consecutively, the findings should be confirmed in platelets from PC, stored for different time periods. In general, further studies are required addressing immune-related mechanisms in stored platelets, including TLR-dependent functions and the influence of storage temperature.The authors have no conflicts of interest to declare.

Abstract lecture

Prof.

Comparative study on the quality of blood products collected and stored in blood bag systems with and without DEHP

Vergleichsstudie über die Qualität von Blutprodukten, die in DEHP-haltigen oder DEHP-freien Blutbeutelsystemen gesammelt und gelagert wurden

Di(2-ethylhexyl) phthalate (DEHP) is an organic compound used to make blood bag systems (BBS) flexible. Concerns about possible endocrine-disruptive effects have led to European legislation aiming to ban the use of these plasticizers. However, dissolved DEHP has a beneficial effect in maintaining red blood cell (RBC) stability. The objective of this study was to collect quality data of blood products stored in BBS with and without DEHP and of RBCs resuspended in different additive solutions.Whole blood was collected and stored in BBS (Fresenius Kabi) with different combinations of plasticizers and RBC additive solutions (A: DEHP-SAGM, B: DEHP-PAGGSM, C: DINCH/BTHC/DEHT-PAGGSM). Collection and processing of the resulting blood products were performed under routine-like conditions. Manufacturing steps were monitored and evaluated. The collection of quality parameters was based on the requirements of the local guidelines and the recommendations of the European Blood Alliance with a strong focus on the quality and shelf-life of RBCs. RBC samples were collected weekly until day 50 after collection. Testing for in vitro quality characteristics was performed according to local standard operation procedures.Leucoreduction filtration times were prolonged in RBCs resuspended in PAGGSM (B: 32.3 ± 6.2 min, C: 34.4 ± 6.8 min) compared to the SAGM group (A: 26.3 ± 4.7 min). Group C (DEHP-free) maintained higher ATP levels during storage time than the other two groups (DEHP, A: 1.88 ± 0.50 µmol/gHb, B: 1.99 ± 0.49 µmol/gHb, C: 2.86 ± 0.53 µmol/gHb). Hemolysis levels were increased in group A compared to group B but comparable to the DEHP-free system (A: 0.24 ± 0.06 %, B: 0.21 ± 0.05 %, C: 0.27 ± 0.10 %). However, no single value exceeded 0.8 % even at day 50. Other parameters, including leucocytes, MCV, pH and Hb, demonstrated only minor differences between the groups. The measured PLT characteristics were unaffected by the bag material.Some differences in RBC parameters were observed between the three combinations of additive solutions and plasticizer types used in the BBS. However, the values of all groups complied with the requirements of the German guidelines even on day 50 after collection. In order to conclusively assess the quality of blood products stored in DEHP-free BBS, it is essential to examine irradiated products and RBCs that are not being mixed during storage due to sampling.This study was supported by Fresenius Kabi (supply with blood bag samples).

Soluble transferrin receptor/Ferritin (sTfR/SF) ratio and haemoglobin concentration outcomes as predictive measure to detect Iron deficiency in frequent blood donations in Nigeria

Blood Donation

Ergebnisse des Verhältnisses von löslichem Transferrinrezeptor/Ferritin (sTfR/SF) und der Hämoglobinkonzentration als prädiktives Maß zur Erkennung von Eisenmangel bei häufigen Blutspenden in Nigeria

Maintaining donor health is of paramount than ensuring a safe, ceaseless blood supply, and blood bays should scout for screening programs for donors to improve their health, recruitment, and withholding.One hundred and eighty-four (184) subjects comprising five groups were recruited into this study, that is, 35 (19.0%) control group, 32 (17.4%) of first, 35 (19.0%) of second, 41 (22.3%) of third and 41 (22.3%) of fourth time blood donors. The donors were within the range of 18 – 49 years of age. Whole blood and serum samples (sera) were used and parameters namely; haemoglobin concentration (Hb) was determined using colorimetric method, serum ferritin level (SF) and serum transferrin receptor level (STfR) were determined using enzyme immunosorbent assay (ELISA), while sTfR was determined by dividing sTfR by SF and finding log of result.Age of donors display that bulk of their ranged 18 -25 in age (45.7%) and 26 – 35 years (47.3%) while ≥ 36 years (7.1%) were very few. 62.5% of 184 blood donors were remunerated while 37.5% were not at the time of donation. Moreover, 26.6% of blood donors donated elsewhere other than the facility while 73.4% donated in the facility. The mean values of Hb and SF decreased significantly (p &lt; 0.001). It was from (152g/L, 142g/L, 125g/L, 115g/L and 103g/L) for the control group to fourth time male donors for Hb, and (101.7, 104.8, 50.1, 33.5 and 21.3 ng/ml) for SF as the number of donation increased within a year. Meanwhile, mean value of sTfR and sTfR/SF ratio from control group to fourth time male blood donors were raised as donation increased significantly (p &lt; 0.01) (2.7, 3.7, 6.6, 8.3 and 10.9 µg/ml) and (1.41, 2.09, 4.76, 8.04 and 14.2 µg/ng) respectively. Male blood donors with iron-deficient as defined by Hb (&lt;125g/L) was 43.5%, SF (&lt;15ng/ml) was 35.3%, sTfR (&gt;3.5µg/ml) was 57.6% and sTfR/SF ratio (&gt;2.5µg/ng) was 56.5% in all 184 male blood donors.Of all the indices used to diagnose iron deficiency, sTfR and sTfR/SF ratio produced the highest percentages of 56.5% and 57.6% respectively of the donors thus, making these indices the most sensitive to diagnose iron deficiency. Blood banks should determine the Hb levels, SF, sTfR and its ratio of blood donors at the time of donor screening to give adequately, the total number of donors having iron deficiency. Also, voluntary non-remuneration should be encouraged.The authors declared no conflicts og Interest

PD Dr. rer. nat.

Prof. Dr.

Universal plasma quality and coagulation parameter stability during storage

Qualitäts- und Gerinnungsparameter von Universalplasma während der Lagerung

Plasma isoagglutinins require ABO blood group compatibility. We have developed a GMP-compliant method to deplete the isoagglutinin titer of fresh frozen plasma (FFP) units of groups A, B and O based on isoagglutinin adsorption on added red cells from red cell concentrates (RCC). We performed an extensive validation study to assess the effect of plasma treatment of blood group A plasma on coagulation and immune parameters directly after manufacture and after storage at ≤30°C for at least 6 months.We have developed a bag system that facilitates the pooling of three units of FFP, addition of the appropriate volume of RCC, and subsequent removal of the isoagglutinin red cell sediment in a closed system. After incubation at 20-24°C for two hours, the isoagglutinin red cell sediment was separated from the plasma pool by centrifugation (4,000g, 10min). We compared plasma units from 12 production processes (totaling 36 units) with and without the addition of RCC for isoagglutinin adsorption, focusing on the following parameters: anti-B titer, hemoglobin, residual erythrocytes, factors II-XIII, fibrinogen, vWF antigen + activity, protein C + S, antithrombin, ADAMTS13, aPTT, Quick value, lactate dehydrogenase, and complement activation immediately after production and after storage at ≤-30 °C for 8.6±1.5 months.From peaks of 1:16 in the plasma pools, total anti-B titers were reduced to &lt; 1:1 in all units in the treatment arm. The coagulation factors II to XIII of the plasma units analyzed were largely within reference ranges even after 6 months of storage, except for factor VIII in both the treatment and control arm. Factor VIII decreased on average by 21.4% (treatment group) and 14.9% (control group) after 6 months of storage, thus fulfilling the requirements of the hemotherapy guidelines. Other as parameters C3 and C4 complement activity, ADAMTS13, INR, vWF antigen and activity and lactate dehydrogenase were within reference ranges after storage. There was no difference between the control and treatment groups.We introduce a fully automated GMP-compliant method for producing universal plasma suitable for blood group-independent transfusion. Our method maintains the quality and safety of universal plasma, as demonstrated by the preservation of all coagulation and immunological parameters even during storage for at least 6 months below -30°C.Macopharma kindly provided the bag systems.

Abstract lecture (Abstract winner)

SARS-CoV-2 convalescent plasma for the controlled clinical trial "COVIC-19": Experience from collection of very high-titer plasma from superimmunized individuals

SARS-CoV-2 Rekonvaleszentenplasma für die klinische Studie "COVIC-19": Erfahrungen mit der Sammlung von sehr hochtitrigem Plasma von superimmunisierten Personen

COVID-19 convalescent plasma (CCP) is a potential therapy for COVID-19. The COVIC-19 trial is building on lessons from previous studies like CAPSID, adopting a novel approach:CCP with very high concentrations of SARS-CoV-2 antibodies (≥4,000 BAU/ml) from donors with previous SARS-CoV-2 infection and vaccinationtreatment of vulnerable persons early after symptom onsetsequencing and cross-neutralization analyses to track viral evolutionWe report the initial experience of collection of very high-titer plasma units for this COVIC-19 trial.We recruited 691 potential donors with previous SARS-CoV-2 infection (inf) and vaccination (vax) who passed initial eligibility check. Anti-SARS-CoV-2 antibody concentrations were measured by anti-SARS-CoV-2 QuantiVac ELISA (Euroimmun) and neutralization capacity was assessed in the GenScript Surrogat and a pseudovirus neutralization assay.We found that high antibody levels correlated with high neutralizing capacity. The average antibody concentration among willing donors was 4,298 BAU/ml. 41.1% of individuals had antibody concentrations of ≥4,000 BAU/ml and concentrations did not significantly differ by gender or ABO type, but were higher among those who had received 3 vax (median 3,746 BAU/ml) or 4 vax (median 4,166 BAU/ml). Highest S-Ab were observed in those with a breakthrough infection after 2 vax, followed by a booster (median 5,159 BAU/ml, 64% ≥4,000 BAU/ml) or breakthrough inf after 3rd or 4th vax (median 3,846; 48% ≥4,000 BAU/ml). Ultimately, 158 eligible individuals donated CCP with concentrations ranging from 4,105 BAU/ml to 22,923 BAU/ml with a median of 7,074 BAU/ml. Majority of donors donated once or twice (35.2% and 20.3%), 13.4% donated 3x, and 31.1% donated ≥4x.Considering all eligibility criteria, 31.9% of the individuals screened met the requirements for high-titer plasma, but only 23% actually donated. Collection of very-high titer plasma from super-immunized individuals with prior infection and vaccination is feasible but requires substantial donor selection, rapid screening and prompt apheresis to capitalize on the short period of very high mAb.COVIC-19 is supported by BMBF.Kein Interessenkonflikt

Hypoxic storage of Leucodepleted Erythrocyte concentrates in PAGGS-M results in increased product quality over conventional products as assessed by Adenosine Triphosphate and 2,3-Diphosphoglycerate

Lagerung von Erythrozytenkonzentraten in Hypoxie verbessert die Qualitätsparamater Adenosin Triphosphat und 2,3-Diphosphoglycerat gegenüber konventionellen Produkten

Storage of Erythrocyte Concentrates (EC) under hypoxia has been made technically possible under GMP. It has been shown to preserve red blood cell (RBC) function, resulting in increased oxygen uptake as well as oxygen release, and extended in vivo survival of transfused RBC. We here aimed to study RBS quality in ECs derived from normal whole blood donations.ECs from 500ml whole blood donations were prepared by the buffy coat method, stored in PAGGS-M solution and de-oxygenated within 24h by flow through CE-marked HemaNext-ONE deoxygenation system (Hemanext, Lexington, MA, USA). They were subsequently stored in oxygen-impermeable bags at 2-6 degrees C over 42 days, and assessed for quality parameters including Adenosin Triphosphate (ATP) and 2,3-Diphosphoglycerate (2,3-DPG).Hypoxia was efficiently reached and maintained between 14 and 17% over the storage time of 42 days. Mean 2,3-DPG levels in deoxygenated RBC amounted to 12 to 16 µmol/g hemoglobin (Hb) between day 0 and day 21 of storage, 6 µmol/g Hb at day 28 and 2 µmol/g Hb at the end of storage, whereas they approached zero values already by day 14 in conventionally stored products. ATP levels were above 4 µmol/g Hb between days 0 and 28 and declined to 3.5 µmol/g Hb at the end of storage at day 42, and were continuously higher compared to conventional RBCs. All other parameters (Hb content, hematocrit, hemloysis) remained within the range of conventional controls.Hypoxic storage of RBSs resulted in significant improvements in red cell strorage quality parameters ATP and 2,3-DPG. Hypoxia-stored RBCs met guideline and license requirements. These findings may also have implications on establishing improved storage conditions of RBCs in Di(2-ethylhexyl)terephthalate (DEHT)-free blood bag systems.Es wurden keine Interessenkonflikte bekannt.

Hypoxic Storage of Leucodepleted Erythrocyte Concentrates in PAGGS-M Results in Increased Product Quality over Conventional Products as Assessed by Adenosine Triphosphate and 2,3-Diphosphoglycerate

Raum 13

Blood donation and components

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Blood donation and components

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