ALBERT

Description: Introduction Cryopreservation for long term storage of platelets (PLTs) represents a clinical useful method for avoiding platelet shortage. Many studies have tried to define, in vitro and in vivo, the entity and weight of storage-related PLTs lesions with discordant results related to different methods. We have performed an in vitro prospective study to evaluate PLTs count, viability and function of buffy coat derived pooled platelet concentrates (BC-PLTs) treated with dimethyl-sulphoxide (DMSO) and cryopreserved at -80°C with an innovative patented system not requiring laminal flow hoods and external manipulations. Materials and methods Each BC-PLTs was obtained from 5 buffy coats and pooled according to standard procedures. The final PLTs concentrates were leukoreduced by filtration and transferred to a 650 mL cryopreservation kit (Promedical ®) which allowed mixing with DMSO 25% in a closed system and following removal of supernatant without further manipulations. BC-PLTs were washed prior freezing with removal of at least 84% supernatant solution, suspended in homologous plasma from 1 of the 5 donors to a final concentration of 200 mL and frozen at – 80°.Selection criteria to make BC-PLTs available for this study was pooled PLTs concentration 〉 1250 x 109/L and a blood units collection time duration shorter than 6 minutes. All the 245 donors were healthy volunteers and they did not take any medication affecting PLTs function. BC-PLTs were analyzed immediately pre-freezing (T0) and 3 months after cryopreservation ( CRY BC-PLTs). The following parameters were assayed: PLTs count (PC), mean platelet volume (MPV), pH, flow cytometry (FACS) expression of CD41a, CD42b, CD61a, CD62p, PAC-1, Annexin V PLTs surface antigens and thromboelastography (TEG). All samples were analyzed also after dilution (1:4) with homologous plasma to approximately 400 x109/L PLTs (data not shown) and for bacterial contamination (BC). CRY BC-PLTs samples were thawed in a bath at 37°C for 5 minutes and evaluated promptly. All the tests were performed according to current European recommendations. PLTs swirl was furthermore visually assessed. Results were expressed as mean +/- standard deviation (SD). Results obtained at T0 and after 3 months were compared by paired sample t-test. Differences were considered as significant at p values

Description: Background: Cryopreservation of platelets (PLTs) at -80°C with dimethyl sulfoxide (DMSO) can extend their shelf life up to 2 years. Cryopreserved PLTs (CRY-PLTs) are reported to have a greater in vivo hemostatic effect than liquid-stored PLTs. Aims of this study were: i. to evaluate the thrombin generation potential of buffy coat derived cryopreserved PLTs (CRY- BC PLT) in comparison with fresh buffy coat derived platelets concentrates; ii. to determine the efficacy and safety of CRY-PLTs transfusion in hematological patients with severe thrombocytopenia. Materials and methods: BC PLTs were obtained from 5 buffy coats and pooled. The final PLTs concentrates were leukoreduced by filtration and transferred to a 650 mL patented cryopreservation kit (Promedical ®) which allowed mixing with DMSO 25% in a closed system and following removal of supernatant without further manipulations. BC-PLTs were washed prior freezing, suspended in homologous plasma from 1 of the 5 donors to a final concentration of 200 mL and frozen at - 80°. CRY- BC PLTs were preserved at -80°C with 6% DMSO. A system of 3 accessory bags directly connected to the mother bag was adopted for the in vitro study, to avoid repeated freezing/thawing of samples. In vitro assays were performed before freezing and at 3,6 and 9 months after thawing. Before assay, CRY-BC PLT were thawed at 37°C and diluted in plasma to adjust to 300× 109/L PLTs. Fresh BC PLTs underwent the same dilution to adjust to 300 ×109/L PLTs. Thrombin generation (TGA) was tested in CRY BC-PLTs and compared to TG potential of fresh BC PLTs. TGA was triggered by the addition of 0.5 pmol/L of recombinant human tissue factor. Endogenous thrombin potential (ETP) and peak height (PH) were determined. Flow Cytometry assays for PLTs activation markers and thromboelastography were also determined on each sample. CRY-BC PLTs, separately prepared according to the above described method for in vivo study, were infused in five hematological patients with acute leukemia (AL) and severe thrombocytopenia (PLTs