Red blood cell preservation by droplet freezing with polyvinylpyrrolidone or sucrose‐dextrose and by bulk freezing with glycerol

Abstract

Background Red blood cell (RBC) preservation is essential to transfusion medicine. Many blood group reference laboratories need a method to preserve rare blood samples for serologic testing at a later date. This study offers a comparison of three common cryoprotective agents and protocols used today: bulk preservation with glycerol and droplet freezing with sucrose/dextrose (S+D) or polyvinyl pyrrolidone (PVP). Study design and methods Human blood from 14 volunteers was collected and frozen at set intervals over two weeks with PVP, S+D, or glycerol. The frozen RBCs were later thawed and the percentage of surviving RBCs was determined. Detailed protocols and an instructional video are supplied. Results Over a two week period, RBCs preserved with glycerol and thawed with a widely used protocol showed a recovery of 41 ± 16 % (mean ± standard deviation) while those thawed with a modified glycerol protocol showed a recovery of 76 ± 8 %. RBCs preserved by droplet freezing with S+D showed a recovery of 56 ± 11 % while those preserved by droplet freezing with PVP showed a recovery of 85 ± 6 %. Recovery values were similar with ethylenediaminetetraacetic acid (EDTA) or heparin anticoagulants, differing freezing rates, and varying droplet volumes. Conclusion Droplet freezing with PVP offered the greatest recovery. While bulk freezing with glycerol can be effective too, droplet freezing may be a more convenient method overall. It requires less effort to thaw, needs much less storage room, and allows blood group laboratories to be frugal with thawing rare samples.

Cryopreservation of glucose-6-phosphate dehydrogenase activity inside red blood cells: developing a specimen repository in support of development and evaluation of glucose-6-phosphate dehydrogenase deficiency tests

“…A glycerol-based protocol for cryopreservation of red blood cells was used [13]. A total of 6 mL of whole blood was transferred to a 15- mL test tube and centrifuged 1,000/g for 5 min.…”

“…The cryovials were removed from −80°C and thawed at room temperature [13]. The red blood cells were transferred to a 5-mL test tube.…”

“…Many blood group reference laboratories have the ability to freeze, reconstitute and analyse red blood cells that were collected months or years earlier [9-11]. Over the years several methods have been used to preserve the metabolic functions of red blood cells [12,13]. In this report, the feasibility of developing a G6PD specimen repository by cryopreservation is evaluated.…”

Incomplete inhibition by eculizumab: mechanistic evidence for residual C5 activity during strong complement activation

“…PNH-RBCs were used immediately or after storage in liquid nitrogen. 36 The fraction of PNH-RBCs type III was determined by fluorescenceactivated cell sorting (FACS) analysis. 37 Sera were collected in VACUETTE/ S-Monovette serum collection tubes, aliquoted, frozen in liquid nitrogen, and stored at 280°C.…”

The Rhesus Site

“…2). Protocols for antibody elution, RBC cryopreservation and DNA extraction as well as flow cytometry are documented; the ‘KryoBeads' or ‘droplet freezing' method is even demonstrated in video clips, which were later revised, published [21] and can be found on YouTube since 2011. The other internal link leads to a comparison of the German D typing recommendations of 1996 and 2000.…”