Human bdnf elisa kit

Patients and healthy controls provided the venous blood samples after a 12 h overnight fast. The blood samples were centrifuged (1200× g, 10 min) within 1 h of collection, and the obtained serum and EDTA-plasma were stored at −80 °C, until further analysis.
Serum fatty acids were determined as methyl esters and separated by gas chromatography. A detailed procedure is described in our most recent previous work [9 (link)]. Omega-6/omega-3 ratio was calculated from (AA + LA)/(EPA + DHA) concentrations in μg/0.1 mL.
Thromboxane B2 in plasma was determined with a Thromboxane B2 EIA kit (Cayman Chemicals, No. 501020, Ann Arbor, Michigan 48108, MI, USA), according to the manufacturer’s protocol. The concentration of thromboxane is presented in pg/mL.
BDNF was determined in plasma with a Human BDNF ELISA kit (Sigma-Aldrich, No. RAB0026, St. Louis, MO, USA), according to the manufacturer’s protocol, and is expressed in ng/mL.
Homocysteine was determined in plasma by Advia Centaur XP HCZ kit (Siemens, Ref 09087913, USA) in the Advia Centaur system (Siemens Healthcare Diagnostics, Tarrytown, NY, USA). The homocysteine level is expressed in µmol/L.
Serum vitamin D was determined by a 25-OH Vitamin D total Elisa kit (Access 2 Immunoassay system, Beckman Coulter, Inc., Brea, CA, USA). The vitamin D concentration is expressed in ng/mL.

To determine levels of 8-OHdG, BDNF and IL-6 in the slice culture medium and the extracts of OBSCs, samples were analyzed using commercially available ELISA kits (New 8-OHdG Check ELISA kit, JaICA Co. Ltd. (Shizuoka, Japan), Human BDNF ELISA kit, RAB0026, Sigma-Aldrich, Mouse IL-6 ELISA Kit, RAB0308-1KT, Sigma-Aldrich, Tokyo, Japan). Culture medium was obtained during and after the cultivation of OBSCs. At the end of cultivation, OBSCs (2 slices) from a single culture membrane were carefully removed from the culture insert without containing matrigel. The OBSCs were then homogenized in 100 μL RIPA lysis buffer (WSE-7420 EzRIPA Lysis kit, ATTO Co., Tokyo, Japan) to obtain the extracts. Protein concentrations of the extracts were measured with a protein quantification kit (Protein Quantification Kit-Rapid, Dojindo Molecular Technologies, Inc., Kumamoto, Japan). For each ELISA, culture medium or OBSCs extracts (with same concentration of protein) were diluted to bring the expected concentration within the range of the standard curve and reacted with first and secondary antibodies, streptavidin-HRP and detection solution according to the manufacturer’s instruction. The reaction was stopped using the stop solution (from the ELISA kits) and absorbance read at 450 nm using a microplate reader (SH-9000Lab, Hitachi, Tokyo, Japan).

Blood samples taken from each participant at the beginning and the end of the study period were analysed by complete blood counts at the Institute of Clinical Chemistry, UKSH Kiel, Kiel, Germany. Additionally, serum samples were used to measure blood concentrations of brain derived neurotrophic factor (BDNF) by using a commercially available Human BDNF ELISA Kit (Sigma-Aldrich, St. Louis, Missouri, US) at the end of the study. The full set of blood measurements is displayed in Suppl. Figure 3. Serum samples were diluted 200-fold and ELISA was performed according to the manufacturers protocol.