Influence of hemolysis, lipemia and bilirubin on biobank sample quality- origin and interference in the use for extracellular vesicle (EV) and MiRNA analyses.

Abstract

Purpose: Pre-analytic interferences can influence the laboratory downstream measurements. We recognized hemolysis, lipemia and bilirubin in some of the serum/plasma samples of the NTF-Biobank from polytraumatized patients. Aim of the present study was to detect interferences, find reasons and describe the influence on downstream analyses.

Methods: The study included serum samples of n = 88 polytraumatized patients admitted to a Level 1 Trauma Center in Germany at the ER & up to 10 days after trauma. Optical absorption spectra of UV-VIS (350-660 nm) were measured to detect hemolysis, lipemia and bilirubin. To find reasons for the interferences, clinical parameters like triglycerides (TAGs), nutrition, anaesthesia or transfusions were collected from patients' record. Extracellular vesicles (EVs) were isolated by SEC from controls, lipidemic and hemolytic samples and analysed via NTA.

Results: Within 10 days after trauma 31.8% of polytraumatized patients' samples showed hemolysis, 12.5% showed increased bilirubin and 15.9% lipemia. Hemolysis occurred in samples mostly at the ER (18%) and was not associated with the number of red blood cell transfusions or the ISS. Both contaminants, hemolysis and lipemia interfered with EV/EV-miRNA measurements. EV miR-16-5p was significantly increased in patients with hemolysis. The presence of lipids further influenced the EV particle size distribution and concentration.

Conclusion: The optical absorption spectra measurement is an easy tool for a robust pre-analytic sample controlling for the presence of interferences. Nutrition and anaesthesia were found to be related with lipemia in samples. Hemolysis and lipemia interfered with EV/EV-miRNA analysis. Therefore, the optical absorption spectra pre-analyses should be incorporated in the EV-biobank sampling.