Quantakine elisa

Venous blood samples were collected in Li-heparin-coated sterile vials (Becton Dickinson, Stockholm, Sweden) and plasma was separated by centrifugation, aliquoted and stored at − 80 °C. Plasma IGF1 was measured using the Human IGF1 ELISA kit (Quantakine ELISA, R&D Systems, Inc., Minneapolis, MN, USA) according to the manufacturer’s instructions. Absorbance was measured at 450 nm (reference 650 nm, wavelength correction set at 540) using a microplate reader and IGF1 concentrations were calculated based on standard curves. The LOD was 0.056 ng/mL, and the coefficient of variation was 5.6%. The plasma concentration of IGFBP3 was measured using Human IGFBP3 solid phase sandwich ELISA (Quantakine ELISA) according to the manufacturer’s instructions. The LOD was 0.14 ng/mL; the coefficient of variance of the assay was 6.2%.

Blood samples were collected and centrifuged for 20 min for 1,500 r.p.m. after coagulation. Serum was stored at −80 °C. Mouse Flt3- ligand Quantakine ELISA was performed according to the manufacturer’s assay procedure (R&D Systems).