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      Effect of Clotting Duration and Temperature on BDNF Measurement in Human Serum

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          Abstract

          Brain-derived neurothrophic factor (BDNF) is a neurotrophin expressed in different tissues and cells, including neurons, endothelial cells, leukocytes, megakaryocytes and platelets. Modifications of BDNF in plasma and/or in serum are associated with neurodegenerative and psychiatric disorders, cardiovascular diseases, metabolic syndrome and with mortality risk. Indeed, changes in blood levels of BDNF may reflect those of its tissue of origin and/or promote pathological dysfunctions. The measurement of BDNF amount in plasma or in serum has been characterized with particular attention in the impact of different anti-coagulants, clotting duration, temperature (≤21 °C) and delay in blood sample centrifugation as well as in stability of storage. However, the influences of normothermic conditions (37 °C) and of clotting duration on BDNF levels in human serum have not been investigated yet. Here, we showed that time and temperature during serum preparation could be taken into consideration to assess the association and/or impact of BDNF levels in the occurrence of pathological conditions.

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          A method for reproducible measurements of serum BDNF: comparison of the performance of six commercial assays

          Alessio Polacchini, Giuliana Metelli, Ruggiero Francavilla … (2015)
          Brain-Derived Neurotrophic Factor (BDNF) has attracted increasing interest as potential biomarker to support the diagnosis or monitor the efficacy of therapies in brain disorders. Circulating BDNF can be measured in serum, plasma or whole blood. However, the use of BDNF as biomarker is limited by the poor reproducibility of results, likely due to the variety of methods used for sample collection and BDNF analysis. To overcome these limitations, using sera from 40 healthy adults, we compared the performance of five ELISA kits (Aviscera-Bioscience, Biosensis, Millipore-ChemiKineTM, Promega-Emax®, R&D-System-Quantikine®) and one multiplexing assay (Millipore-Milliplex®). All kits showed 100% sample recovery and comparable range. However, they exhibited very different inter-assay variations from 5% to 20%. Inter-assay variations were higher than those declared by the manufacturers with only one exception which also had the best overall performance. Dot-blot analysis revealed that two kits selectively recognize mature BDNF, while the others reacted with both pro-BDNF and mature BDNF. In conclusion, we identified two assays to obtain reliable measurements of human serum BDNF, suitable for future clinical applications.
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            Circulating Brain-Derived Neurotrophic Factor and Indices of Metabolic and Cardiovascular Health: Data from the Baltimore Longitudinal Study of Aging

            Erin Golden, Ana B F Emiliano, Stuart Maudsley … (2010)
            Background Besides its well-established role in nerve cell survival and adaptive plasticity, brain-derived neurotrophic factor (BDNF) is also involved in energy homeostasis and cardiovascular regulation. Although BDNF is present in the systemic circulation, it is unknown whether plasma BDNF correlates with circulating markers of dysregulated metabolism and an adverse cardiovascular profile. Methodology/Principal Findings To determine whether circulating BDNF correlates with indices of metabolic and cardiovascular health, we measured plasma BDNF levels in 496 middle-age and elderly subjects (mean age ∼70), in the Baltimore Longitudinal Study of Aging. Linear regression analysis revealed that plasma BDNF is associated with risk factors for cardiovascular disease and metabolic syndrome, regardless of age. In females, BDNF was positively correlated with BMI, fat mass, diastolic blood pressure, total cholesterol, and LDL-cholesterol, and inversely correlated with folate. In males, BDNF was positively correlated with diastolic blood pressure, triglycerides, free thiiodo-thyronine (FT3), and bioavailable testosterone, and inversely correlated with sex-hormone binding globulin, and adiponectin. Conclusion/Significance Plasma BDNF significantly correlates with multiple risk factors for metabolic syndrome and cardiovascular dysfunction. Whether BDNF contributes to the pathogenesis of these disorders or functions in adaptive responses to cellular stress (as occurs in the brain) remains to be determined.
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              Measurements of brain-derived neurotrophic factor: methodological aspects and demographical data.

              B Marcussen, Per Hartvig Honoré, Maj Vinberg … (2007)
              Although numerous studies have dealt with changes in blood brain-derived neurotrophic factor (BDNF), methodological issues about BDNF measurements have only been incompletely resolved. We validated BDNF ELISA with respect to accuracy, reproducibility and the effect of storage and repeated freezing cycles on BDNF concentrations. Additionally, the effect of demographic characteristics in healthy subjects on BDNF was verified. Whole blood and serum was collected from 206 healthy subjects and a subgroup was genotyped for BDNF Val66Met polymorphism. The effect of age, gender, BDNF genotype and thrombocyte count on whole blood BDNF was assessed. The BDNF ELISA measurement was accurate, 91.6+/-3.0%, and showed high reproducibility, whereas inter-assay and intra-subject variations were modest, 8.4+/-5.2% and 17.5+/-14.1%, respectively. Storage of whole blood samples at 4 degrees C significantly decreased BDNF concentration, while repeated freezing cycles and storage at -20 degrees C was without any effect. Storage at -20 degrees C of serum, but not whole blood, was associated with a significant decrease in BDNF concentration. Women had significantly higher whole blood BDNF concentrations than men (18.6+/-1.3 ng/ml versus 16.5+/-1.4 ng/ml), and showed a right-skewed BDNF concentration distribution. No association between whole blood BDNF concentrations and thrombocyte count, age, or BDNF genotype was found. In conclusion, the BDNF ELISA assay determines whole blood BDNF accurately and with high reproducibility. Female gender is associated with higher whole blood BDNF concentrations whereas age, thrombocyte count and BDNF Val66Met polymorphism were un-associated.
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                Author and article information

                Journal
                Journal ID (nlm-ta): Int J Mol Sci
                Journal ID (iso-abbrev): Int J Mol Sci
                Journal ID (publisher-id): ijms
                Title: International Journal of Molecular Sciences
                Publisher: MDPI
                ISSN (Electronic): 1422-0067
                Publication date (Electronic): 15 September 2017
                Publication date Collection: September 2017
                Volume: 18
                Issue: 9
                Electronic Location Identifier: 1987
                Affiliations
                [1 ]Centro Cardiologico Monzino, IRCCS, 20138 Milan, Italy; patrizia.amadio@ 123456ccfm.it (P.A.); leonardo.sandrini@ 123456unimi.it (L.S.); elena.tremoli@ 123456ccfm.it (E.T.)
                [2 ]Department of Pharmacological and Biomolecular Sciences, University of Milan, 20133 Milan, Italy; alessandro.ieraci@ 123456unimi.it
                Author notes
                [* ]Correspondence: silvia.barbieri@ 123456ccfm.it ; Tel.: +39-02-5800-2021
                Author information
                https://orcid.org/0000-0001-6737-7695
                https://orcid.org/0000-0002-7486-2637
                Article
                Publisher ID: ijms-18-01987
                DOI: 10.3390/ijms18091987
                PMC ID: 5618636
                PubMed ID: 28914800
                SO-VID: 0db876e7-83fe-4f6d-9022-05aebf0f0e30
                Copyright © © 2017 by the authors.
                License:

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                Date received : 11 August 2017
                Date accepted : 02 September 2017
                Categories
                Subject: Communication

                ScienceOpen disciplines: Molecular biology
                Keywords: bdnf,temperature,clotting duration,serum
                Data availability:
                ScienceOpen disciplines: Molecular biology
                Keywords: bdnf, temperature, clotting duration, serum

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