Human igf 1 elisa kit

Blood samples were drawn from all ISSHL participants from the median cubital vein upon enrolment, and on Day 21, samples were centrifuged to obtain the serum fraction which was stored at −80°C until analysis. Serum levels of IGF-1 and HSP70 were determined using a Human IGF-1 ELISA kit and a Human HSP70 ELISA kit (R&D Systems, Minneapolis, MN, USA), respectively, according to the manufacturer's protocols. The change of IGF-1(△IGF-1)=(post-therapy IGF-1) − (initial IGF-1) and the change of HSP70(△HSP70) = (post-therapy HSP70) − (initial HSP70).

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%.

A human IGF-1 ELISA kit (R&D System Inc., Minneapolis, MN, USA) was used to measure rhIGF-1 levels. The rhIGF-1 solution (6 ng/mL to 0.1 ng/mL) was serially diluted for each set of perilymph samples to produce a standard concentration curve. The test and control samples, as well as the positive and negative controls provided by the manufacturer, were added to a 96-well microplate and incubated with the diluted drug-enzyme conjugate at room temperature for 1 h. The plate was then washed 3 times with diluted wash buffer to remove any unbound sample or drug-enzyme conjugate. Substrate solution was added and incubated for 30 min. The reaction was then halted with stop solution. The plate was read with an ELISA microplate reader equipped with a 450-nm filter (Synergy H4 Hybrid Reader). The absorbance value obtained by the ELISA reader was plotted and compared to the standard concentration curve to calculate the concentration of rhIGF- 1 present in each perilymph sample. In this assay, the minimal detectable perilymph concentration of rhIGF-1 was 0.1 ng/mL.