Freezing of buffy coat–derived, leukoreduced platelet concentrates in 6 percent dimethyl sulfoxide

Abstract

There has recently been renewed interest in freezing platelets (PLTs) in dimethyl sulfoxide (DMSO) for the treatment of major traumatic injuries, especially in military situations. This study examined PLTs that were frozen in small volumes of 6 percent DMSO at -80 degrees C. Buffy coat-derived pooled leukoreduced PLT concentrates were frozen in 6 percent DMSO and stored at -80 degrees C. Assays included hypotonic shock response (HSR); aggregation; glycoprotein (GP)Ibalpha and P-selectin binding sites; annexin V binding to phosphatidylserine, glycocalicin, and lactate dehydrogenase (LDH). Cone and plate technology (DiaMed Impact-R, DiaMed) was used to test PLT function under near physiologic conditions. The freeze-thaw loss of PLTs was 23 percent. HSR was 17 +/- 7 percent. Cytometry demonstrated two populations of PLTs: one with normal levels of GPIbalpha binding sites (27 x 10(3) +/- 3 x 10(3)/PLT) and one with reduced levels (5.5 x 10(3) +/- 1.2 x 10(3)/PLT). There were 1.4 x 10(3) +/- 0.2 x 10(3) P-selectin binding sites per PLT. Annexin V binding to phosphatidylserine was 50 +/- 9 percent and LDH was 496 +/- 207 IU per 10(12) PLTs. Surface coverage and aggregate size, as measured by the DiaMed Impact-R, were similar to those observed with PLTs stored for 2 days at 22 degrees C. Some degree of activation was demonstrated by the proportion of PLTs with reduced levels of GPIbalpha binding sites, increased P-selectin expression, and increased Annexin V binding. LDH concentrations indicated a degree of lysis. The DiaMed Impact-R results showed that the PLTs were still capable of adhering to surfaces and forming aggregates under shear force.