Abstract
Circulating concentrations of brain-derived neurotrophic factor (BDNF) have been linked to cancer, neuropsychiatric, diabetes, and gynecological disorders. However, factors influencing plasma storage and subsequent BDNF quantification are incompletely understood. Therefore, the anticoagulant used in plasma separator tubes, storage-time, storage-temperature, and repeated freeze–thaw cycles on circulating BDNF concentrations was evaluated. Peripheral blood samples were collected from healthy women (n = 14) and men (n = 10) recruited prospectively from McMaster University (August 2014). Blood was collected from the cubital vein into plasma separator tubes containing five different anticoagulant systems [K2EDTA, Li-Hep, Li-Hep (gel), Na-Hep, Na-Hep (glass)], and placed on ice for transport to the lab for centrifugation. Plasma samples (n = 16) collected in K2EDTA tubes from women recruited to a previous study (April 2011 to December 2012) were used to determine the effect of multiple freeze–thaw cycles. Plasma BDNF was quantified using a commercially available ELISA kit. Plasma concentrations of BDNF were significantly affected by the type of plasma separator tube, storage-time, and number of freeze–thaw cycles. Storage temperature (− 20 vs. − 80 °C) did not significantly affect the quantity of BDNF measured as mean BDNF concentrations generally fell within our calculated acceptable change limit up to 6 months in the freezer. Our results suggest that for quantification of circulating BDNF blood collected in K2EDTA tubes and plasma stored up to 6 months at either − 20 or − 80 °C produces reproducible results that fall within an acceptable range. However, plasma samples stored beyond 6 months and repeated freeze–thaw cycles should be avoided.
Highlights
Circulating concentrations of brain-derived neurotrophic factor (BDNF) have been linked to cancer, neuropsychiatric, diabetes, and gynecological disorders
Plasma samples collected between April 2011 and December 2012 in K2EDTA tubes from women (n = 16) in a prior study[20] according to our approved ethics protocol (REB#06-064) were available to determine the effect of multiple freeze–thaw cycles on BDNF concentration, as described below
We demonstrate that freezer storage temperature (− 20 oC vs. − 80 oC) does not significantly impact the amount of BDNF recovered from frozen plasma, plasma stored at − 80 °C tended to have less variability in mean BDNF concentrations between time points
Summary
Circulating concentrations of brain-derived neurotrophic factor (BDNF) have been linked to cancer, neuropsychiatric, diabetes, and gynecological disorders. The anticoagulant used in plasma separator tubes, storage-time, storage-temperature, and repeated freeze–thaw cycles on circulating BDNF concentrations was evaluated. Plasma concentrations of BDNF were significantly affected by the type of plasma separator tube, storage-time, and number of freeze–thaw cycles. Our results suggest that for quantification of circulating BDNF blood collected in K2EDTA tubes and plasma stored up to 6 months at either − 20 or − 80 °C produces reproducible results that fall within an acceptable range. The effect of the type of anticoagulant used during plasma collection, sample handling, processing, storage, and repeated freeze–thaw cycles on BDNF concentration are ill defined, rendering comparison of BDNF values between studies difficult. The objective of this study was to evaluate the effect of anticoagulant routinely used in commercial plasma collection tubes, freezer storage-temperature, storage-time, and repeated freeze–thaw cycles on plasma concentrations of BDNF
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