Impact of technical and assay variation on reporting of hemolysis in stored red blood cell products.

BACKGROUND: Hemolysis of RBCs is an important measure of product quality and is influenced by donor factors, blood component manufacturing and storage. Percent hemolysis is determined using hematocrit (Hct), supernatant Hb (SHb) and total Hb (THb), each of which can be measured using a variety of methods.

METHODS: Sixteen members of an international collaborative were surveyed to understand equipment and procedural variation in hemolysis testing. In a laboratory-based evaluation, we examined how hemolysis was impacted by: measurement of Hct, SHb, THb and number and force of centrifugations for SHb preparation. The number and size of extracellular vesicles (EVs) was also examined.

RESULTS: There was no consensus in equipment or procedures used by international laboratories to measure hemolysis. The centrifugation force used to prepare samples influenced SHb concentration when a single or double (p=0.0001) centrifugation step was used. The number and force of centrifugation related directly to the ability to remove EVs and EV-bound Hb from samples. Hemolysis varied significantly from 0.16% to 0.32% (mean of 0.22%) depending on the combination of methods or centrifugation conditions used to test expired samples.

CONCLUSION: Method and preparative procedures have a critical impact on measurement of hemolysis in RCC.