Cryopreservation of buffy coat derived platelets: Paired in vitro characterization using uncontrolled versus controlled freezing rate protocols

白膜血小板的冷冻保存:使用不受控与受控冷冻速率方案进行配对体外表征

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作者:Nahreen Tynngård, Alice Bell, Gunilla Gryfelt, Stefan Cvetkovic, Agneta Wikman, Michael Uhlin, Per Sandgren

Background

Cryopreserved platelets show a reduced recovery and viability after freezing and thawing including several ultrastructural and phenotypic deteriorations compared with liquid-stored platelets. It is suggested that using Controlled-Rate Freezing (CRF) can reduce variability and optimize the functionality profile for cells. The

Conclusion

The use of a uncontrolled freezing rate protocol is feasible, creating a platelet product comparable to using a controlled rate freezing equipment during cryopreservation of platelets.

Methods

To evaluate the possible effects of different freezing rate protocols a two-experimental study comparing diverse combinations was tested with a pool and split design. Uncontrolled freezing of platelets in materials with different thermal conductivity (metal vs cardboard) was evaluated in experiment 1. Experiment 2 evaluated uncontrolled vs a controlled-rate freezing protocol in metal boxes. All variables were assessed pre and post cryopreservation.

Results

Directly after thawing, no major differences in platelet recovery, LDH, ATP, Δψ, CD62P, CD42b, platelet endothelial cell adhesion molecule and sCD40L were seen between units frozen with different thermal conductivity for temperature. In contrast, we observed signs of increased activation after freezing using the CRF protocol, reflected by increased cell surface expression of CD62P, PAC-1 binding and increased concentration of LDH. Agonist induced expression of a conformational epitope on the GPIIb/IIIa complex and contribution to blood coagulation in an experimental rotational thromboelastometry setup were not statistically different between the groups.

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