Thomas Müller

Large clinical trials have demonstrated that some patient groups with hypoproliferative thrombocytopenia benefit from prophylactic platelet transfusions, while in others, a therapeutic transfusion regimen might be sufficient. The remaining capacity to generate endogenous platelets might be helpful to select the platelet transfusion regimen. We assessed whether the recently described method of digital droplet polymerase chain reaction (PCR) can be used to assess the endogenous platelet levels in two groups of patients undergoing high-dose chemotherapy with autologous stem cell transplantation (ASCT). Multiple myeloma (n = 22) patients received high-dose melphalan alone (HDMA); lymphoma patients (n = 15) received BEAM or TEAM (B/TEAM) conditioning. Patients with a total platelet count + -cell dose in the graft was inversely correlated with the intensity of endogenous thrombocytopenia in B/TEAM-treated patients. Monitoring endogenous platelet counts detects the direct effects of myelosuppressive chemotherapies on platelet regeneration. This approach may help to develop a platelet transfusion regimen tailored to specific patient groups.

Background and Objectives: Corrected count increment (CCI) measurements monitor the effectiveness of platelet transfusions in haemato‐oncology, but they usually fail in patients undergoing cardiac surgery. We investigated whether polymerase chain reaction (PCR) of mitochondrial single‐nucleotide polymorphisms (SNPs) is able to monitor the survival of transfused platelets in these patients.Materials and Methods: Leukocyte‐free, platelet‐rich plasma was prepared from patients' blood to measure platelet counts based on patient‐/donor‐specific SNPs by digital PCR after DNA extraction. Platelet counts in samples from patients with severe thrombocytopenia were analysed by both PCR and flow cytometry. Ten patients undergoing cardiac surgery with the use of heart lung machine and without overt bleeding received a single apheresis platelet concentrate because of either dual platelet inhibition during a non‐elective intervention or a complex procedure. Blood samples were collected at nine defined intervals (0–120 h) post transfusion.Results: The digital PCR of the seven SNPs reliably quantified levels ≥0.6 G/L platelets, in good agreement with flow cytometry and without interference by other SNPs or by platelet activation. A mean 24‐h CCI of 11.8 (range: 5.6–19.8) and a mean 120‐h area under the curve (AUC) of 1386 (915–1821) hxG/L were observed for the transfused platelets. The mean AUC of 14,103 (3415–27,305) hxG/L for the patients' endogenous platelets indicates that transfused platelets represented only 11% (5–25) of the total platelet counts during 120 h post transfusion.Conclusion: PCR of mitochondrial SNPs offers a tool to assess the survival of platelets from apheresis concentrates in cardiac surgery patients to facilitate the implementation of improved transfusion strategies.

Background: Emerging viruses like severe acute respiratory syndrome coronavirus (SARS‐CoV), Crimean–Congo haemorrhagic fever virus (CCHFV) and Nipah virus (NiV) have been identified to pose a potential threat to transfusion safety. In this study, the ability of the THERAFLEX UV‐Platelets and THERAFLEX MB‐Plasma pathogen inactivation systems to inactivate these viruses in platelet concentrates and plasma, respectively, was investigated.Materials and methods: Blood products were spiked with SARS‐CoV, CCHFV or NiV, and then treated with increasing doses of UVC light (THERAFLEX UV‐Platelets) or with methylene blue (MB) plus increasing doses of visible light (MB/light; THERAFLEX MB‐Plasma). Samples were taken before and after treatment with each illumination dose and tested for residual infectivity.Results: Treatment with half to three‐fourths of the full UVC dose (0·2 J/cm²) reduced the infectivity of SARS‐CoV (≥3·4 log), CCHFV (≥2·2 log) and NiV (≥4·3 log) to the limit of detection (LOD) in platelet concentrates, and treatment with MB and a fourth of the full light dose (120 J/cm²) decreased that of SARS‐CoV (≥3·1 log), CCHFV (≥3·2 log) and NiV (≥2·7 log) to the LOD in plasma.Conclusion: Our study demonstrates that both THERAFLEX UV‐Platelets (UVC) and THERAFLEX MB‐Plasma (MB/light) effectively reduce the infectivity of SARS‐CoV, CCHFV and NiV in platelet concentrates and plasma, respectively.

BACKGROUND: The THERAFLEX UV‐Platelets system (Maco Pharma) uses ultraviolet C (UVC) light for pathogen inactivation (PI) of platelet concentrates (PCs) without any additional photoactive compound. The aim of the study was to systematically investigate bacterial inactivation with this system under conditions of intended use.STUDY DESIGN AND METHODS: The robustness of the system was evaluated by assessing its capacity to inactivate high concentrations of different bacterial species in accordance with World Health Organization guidelines. The optimal use of the PI system was explored in time‐to‐treatment experiments by testing its ability to sterilize PCs contaminated with low levels of bacteria on the day of manufacture (target concentration, 100 colony‐forming units/unit). The bacteria panel used for spiking experiments in this study included the World Health Organization International Repository Platelet Transfusion Relevant Reference Strains (n = 14), commercially available strains (n = 13), and in‐house clinical isolates (n = 2).RESULTS: Mean log reduction factors after UVC treatment ranged from 3.1 to 7.5 and varied between different strains of the same species. All PCs (n = 12/species) spiked with up to 200 colony‐forming units/bag remained sterile until the end of storage when UVC treated 6 hours after spiking. UVC treatment 8 hours after spiking resulted in single breakthrough contaminations with the fast‐growing species Escherichia coli and Streptococcus pyogenes.CONCLUSION: The UVC‐based THERAFLEX UV‐Platelets system efficiently inactivates transfusion‐relevant bacterial species in PCs. The comprehensive data from this study may provide a valuable basis for the optimal use of this UVC‐based PI system.

BACKGROUND: Nonenveloped transfusion‐transmissible viruses such as hepatitis A virus (HAV) and hepatitis E virus (HEV) are resistant to many of the common virus inactivation procedures for blood products. This study investigated the pathogen inactivation (PI) efficacy of the THERAFLEX UV‐Platelets system against two nonenveloped viruses: HAV and feline calicivirus (FCV), in platelet concentrates (PCs). STUDY DESIGN AND METHODS: PCs in additive solution were spiked with high titers of cell culture–derived HAV and FCV, and treated with ultraviolet C at various doses. Pre‐ and posttreatment samples were taken and the level of viral infectivity determined at each dose. For some samples, large‐volume plating was performed to improve the detection limit of the virus assay. RESULTS: THERAFLEX UV‐Platelets reduced HAV titers in PCs to the limit of detection, resulting in a virus reduction factor of greater than 4.2 log steps, and reduced FCV infectivity in PCs by 3.0 ± 0.2 log steps. CONCLUSIONS: THERAFLEX UV‐Platelets effectively inactivates HAV and FCV in platelet units.

Background: PCR with sequence-specific priming using allele-specific fluorescent primers and analysis on a capillary sequencer is a standard technique for DNA typing. We aimed to adapt this method for donor typing in a medium-throughput setting. Methods: Using the Extract-N-Amp PCR system, we devised a set of eight multiplex allele-specific PCR with fluorescent primers for Fy^a/Fy^b, Jk^a/Jk^b, M/N, and S/s. The alleles of a gene were discriminated by the fluorescent color; donor and polymorphism investigated were encoded by product length. Time, cost, and routine performance were collated. Discrepant samples were investigated by sequencing. The association of new alleles with the phenotype was evaluated by a step-wise statistical analysis. Results: On validation using 312 samples, for 1.1% of antigens (in 5.4% of samples) no prediction was obtained. 99.96% of predictions were correct. Consumable cost per donor were below EUR 2.00. In routine use, 92.2% of samples were successfully predicted. Discrepancies were most frequently due to technical reasons. A total of 11 previously unknown alleles were detected in the Kell, Lutheran, and Colton blood group systems. In 2017, more than 20% of the RBC units prepared by our institution were from donors with predicted antigen status. A steady supply of Yt(a-), Co(a-) and Lu(b-) RBC units was ensured. Conclusions: Pooled capillary electrophoresis offers a suitable alternative to other methods for extended donor DNA typing. Establishing a large cohort of typed donors improved transfusion support for patients.