Human ifn gamma uncoated elisa

The effect of cytokine release on the target cells (K562) was measured using the enzyme-linked immunosorbent assay (ELISA) method. The supernatant collected from the co-culture of the tumor cell media and NK cells was used to investigate the concentration of IFN-gamma and TNF-alpha using Invitrogen's ELISA kit (Human IFN-gamma uncoated ELISA, Human TNF-alpha uncoated ELISA), as per the manufacturer's instructions.

The concentration of TNFα, IL1β, IL10, CXCL10, IL17, and IFNγ in the cell supernatants was measured using an enzyme-linked immunosorbent assay (ELISA) according to the manufacturers’ protocols (Human TNFα Uncoated ELISA, eBioscience, San Diego, CA; Human IL1β Uncoated ELISA, eBioscience; ELISA MAX Deluxe Set Human IL10, BioLegend, San Diego, CA; ELISA MAX Deluxe Set Human CXCL10 (IP-10), BioLegend; Human IL17A alpha Uncoated ELISA, Invitrogen; Human IFN gamma Uncoated ELISA, Invitrogen, Waltham, MA). LPS concentration in the FS samples was assessed by the LPS ELISA Kit (Aviva Systems Biology, San Diego, CA). The concentration of calprotectin in fecal samples was determined by a sandwich ELISA (Calprotectin ELISA; Bühlmann Laboratories AG, Basel, Switzerland).

To investigate the functionality of NK92 cells encapsulated in a porous hydrogel, crosslinked alginate in the hydrogel was degraded using 8 mM Ethylenediaminetetraaceticacid (EDTA) (Supplementary Fig. 3) [29 (link)]. The hydrogel was incubated for 30 min and centrifuged to obtain 3D cultured cells. The cytotoxicity was investigated using a calcein-AM-based assay. Target cancer cells were stained with calcein-AM (Invitrogen, USA) for 1 h at 37 °C. The labeled target cells (1 × 10⁴ cells / 100 µL) and NK92 cells were placed in 96-well round-bottom plates at effector-to-target (E/T) ratios of 2:1 and 1:1 for 4 h. The maximum release was measured by adding 4% Triton X-100 to the target cell. The percentage cytotoxicity was calculated using the following formula: (sample release – spontaneous release) / (maximum release – spontaneous release) × 100.
Cytokine release, which affects the target, was measured using an ELISA. The supernatant of the tumor cell media co-cultured with NK cells was collected, and the concentration of released cytokines (interferon gamma, TNF-alpha) was investigated. An ELISA kit (Human IFN gamma uncoated ELISA, Human TNF alpha uncoated ELISA; Invitrogen, USA) was used according to the instructions of the manufacturer.