Human perforin elisa kit

Granzyme B and perforin secreted from NK cells were investigated under coculture conditions with K562 tumor cells. NK cells were seeded at 40,000 cells per well and cocultured with 4,000 K562 cells per well for 3 hours. The concentrations of lytic granules were determined using a granzyme B ELISA kit (Mabtech, Cincinnati, OH, USA) and a human perforin ELISA kit (Abcam, Cambridge, MA, USA). After the experiments were performed according to the manufacturer's instructions, the results were read using a spectrophotometer (Molecular Devices), and the concentrations of granules were analyzed using GraphPad Prism 8 software.

Perforin secretion was measured using Human Perforin ELISA kit from Abcam (Cambridge. MA). Human interferon gamma (IFN-γ) and tumor necrosis factor (TNF-α) secretion by NK cells were measured by Quantikine ELISA kits (R&D Systems, Minneapolis, MN, USA) as per manufacturer’s instructions. Conditioned media (CM) was collected from NK cells maintained at a concentration 1 × 10⁶ cells/mL, or NK and SUDHL10 cell mixed cultures where each cell type was kept at 0.5 × 10⁶ cells/mL. Cell debris was removed by centrifugation. Briefly, 100 µL of standard IFN-γ, TNF-α, or perforin control or cell culture supernatant were added to the antibody-coated microplate strips and incubated at room temperature for 2 h. The wells were washed thrice with the assay buffer. 200 µL of horseradish peroxidase-conjugated antibodies were added to each well for further incubation of 2 h. The wells were washed repeatedly and incubated with a combination of hydrogen peroxide and tetramethylbenzidine for 30 min. The reaction was stopped with 2N sulfuric acid and the plate read at 450 nm with a correction readout set to 540 nm.

Perforin expression in conditioned medium, mouse serum and harvested xenograft tumors was measured by human perforin ELISA kit according to manufacturer’s instruction (Abcam, Cambridge, UK).