Il 6 human uncoated elisa kit

The effect of the HA nonwoven textiles on immune cell activation was assessed using the Monocyte Activation Test (MAT). The textile samples were dissolved in normal saline (0.9% NaCl) at a concentration of 1 mg/mL. Heparinised whole blood from four healthy donors was pooled and 10× diluted with normal saline. The diluted blood was placed in sterile microtubes and 100 µL of the sample solution was added, followed by incubation at 37 °C for 16 to 20 h. In parallel, the inner control sample was prepared by further adding Reference Standard Endotoxin (RSE, Merck, Darmstadt, Germany, cat. no. #E0150000) at a concentration of 0.25 EU/mL to the sample solution. The purpose of the inner control sample was to exclude undesired interaction between the sample and the RSE lipopolysaccharide. For evaluation purposes, also solutions of pure RSE at concentrations of 10, 5, 1, 0.25 and 0.1 EU/mL were tested.
After incubation, the microtubes were manually shaken and centrifuged at 13 000 rpm for 10 min. The upper phase (plasma diluted in the saline) was collected for IL-6 analysis using an IL-6 Human Uncoated ELISA kit (Invitrogen, Waltham, MA, USA, #88-7066-88) according to the manufacturer’s protocol. Absorbance was read by an EnSight Multimode Reader and the Kaleido software (both from PerkinElmer, Waltham, MA, USA) was used for evaluation.

The levels of IL‐6, IL‐10, and TNF‐α in the supernatants were determined using the following　commercial enzyme‐linked immunosorbent assay (ELISA) kits according to the manufacturer's protocol; IL‐6 Human Uncoated ELISA Kit (88‐7066‐88), IL‐10 Human Uncoated ELISA Kit (88‐7106‐88), and TNF alpha Human Uncoated ELISA Kit (88‐7346‐88) (all from Invitrogen).

Cell-free supernatants from HAF, transfected with either the EGFP or Piezo1 expression plasmid, were collected just before the start of nanoindentation experiments (3 days post-transfection) and stored at −20 °C. IL-6 was detected (in Nunc MaxiSorp 96-well plates (11530627, Invitrogen, Karlsruhe, Germany)) using a colorimetric assay based on the IL-6 Human Uncoated ELISA Kit (88-7066-88, Invitrogen) according to manufacturer’s instructions. Absorbance was measured with a Microplate Reader (Tecan Infinite 200) and analyzed with Magellan data analysis software.

Cytokine production was analyzed in the supernatant of MNCs before and after cryopreservation. Reactiveness of the immune response to a bacterial stimulus was assessed by stimulating the cells with LPS as stated above. Secreted protein amounts were quantified using the TNF alpha Human Uncoated ELISA Kit with plates as well as the IL-6 Human Uncoated ELISA Kit (both ThermoFisher, Waltham, MA, USA) as indicated by the manufacturers’ instructions.

Type I collagen was measured in cell culture supernatants using Human Procollagen I alpha 1 DuoSet ELISA (R&D Systems, Minneapolis, USA) and IL-6 was measured in cell culture supernatants using IL-6 Human Uncoated ELISA Kit (Thermo Fisher Scientific) according to the manufacturer’s protocol.

The concentrations of interleukin (IL-)6 and IL-8 were determined in supernatants of osteoblastic cell cultures. The IL-6 Human uncoated ELISA Kit and the IL-8 Human uncoated ELISA Kit (both: Thermo Fisher Scientific, Waltham, MA, USA) were implemented as described in the manufacturer’s instructions. The absorption was measured using an Infinite F200 pro (Tecan Trading AG, Maennedorf, Switzerland) at a wavelength of 405 nm. The measured IL-6 and IL-8 concentrations were normalized using the corresponding total protein content of the supernatant.

Interleukin 6 (IL6) secretion levels were measured in culture supernatants of HDF cells, by ELISA using the IL6 Human Uncoated ELISA Kit (88-7066-88, Thermo Fisher Scientific) following the manufacturer’s protocol.