Human ifng quantikine elisa kit

Manufactured by R&D Systems

The Human IFNg Quantikine ELISA kit is a quantitative sandwich enzyme immunoassay designed for the measurement of human interferon gamma (IFNg) concentrations in cell culture supernatants, serum, and plasma. The kit utilizes a microplate pre-coated with a monoclonal antibody specific for IFNg. Samples and standards are pipetted into the wells, and any IFNg present is bound by the immobilized antibody. After washing, an enzyme-linked polyclonal antibody specific for IFNg is added to the wells. Following a wash to remove any unbound antibody-enzyme reagent, a substrate solution is added, and color develops in proportion to the amount of IFNg bound. The optical density of the color is then measured.

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Lab products found in correlation

Easysep human nk enrichment kit, by STEMCELL (2 mentions) Lsrii flow cytometer, by Tree Star (1 mentions) Live dead near ir viability stain, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

Quantikine ELISA kit Quantikine ELISA Quantikine Human Quantikine kits Quantikines Human kits ELISAs

4 protocols using human ifng quantikine elisa kit

NK Cell Isolation and Stimulation

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Example 8

Materials and Methods

NK cells were isolated from pooled splenocytes of SRG or SRG-15 mice (3 spleens per group) and NK cells were isolated using EasySep Human NK enrichment kit (StemCell Technologies; Cat #19055).

NK cells were also isolated from healthy human PBMCs. NK cells were treated overnight with 10 ng/mL human IL-2. The next day, cells were stimulated overnight with 10 ng/mL human IL-12p70 or 2 mg/mL poly I:C or left untreated. The next day, supernatant was harvested and IFNg levels assessed using Human IFNg Quantikine ELISA kit (R&D systems; Cat # DIF50). NK cell purity was analyzed by FACS and IFNg levels normalized as picograms (pg) produced by individual NK cells. Statistical analysis was performed using ANOVA test.

Results

As shown in FIG. 24B, SRG and SRG-15 derived NK cells have comparable IFNγ secretion, but less than human PBMC-derived NK cells upon IL-12p70 treatment.

US10561126B2. Genetically modified non-human animals and methods of use thereof (2020-02-18). Regeneron Pharmaceuticals, Inc. [US], Yale University [US], Institute for Research in Biomedicine (IRB) [CH]. Inventors: Dietmar Herndler-Brandstetter [US], Richard A. Flavell [US], Davor Frleta [US], Cagan Gurer [US], Markus Gabriel Manz [CH], Andrew J. Murphy [US], Noah W. Palm [US], Liang Shan [US], Sean Stevens [US], Till Strowig [DE], George D. Yancopoulos [US], Marcel de Zoete [NL].

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Comparative Analysis of NK Cell IFNγ Secretion

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Example 8

Materials and Methods

NK cells were isolated from pooled splenocytes of SRG or SRG-15 mice (3 spleens per group) and NK cells were isolated using EasySep Human NK enrichment kit (StemCell Technologies; Cat #19055).

NK cells were also isolated from healthy human PBMCs. NK cells were treated overnight with 10 ng/mL human IL-2. The next day, cells were stimulated overnight with 10 ng/mL human IL-12p70 or 2 mg/mL poly I:C or left untreated. The next day, supernatant was harvested and IFNg levels assessed using Human IFNg Quantikine ELISA kit (R&D systems; Cat #DIF50). NK cell purity was analyzed by FACS and IFNg levels normalized as picograms (pg) produced by individual NK cells. Statistical analysis was performed using ANOVA test.

Results

As shown in FIG. 24B, SRG and SRG-15 derived NK cells have comparable IFNγ secretion, but less than human PBMC-derived NK cells upon IL-12p70 treatment.

US11576356B2. Genetically modified non-human animals and methods of use thereof (2023-02-14). Regeneren Pharmaceuticals, Inc. [US], Yale University [US], Institute for Research in Biomedicine (IRB) [CH]. Inventors: Dietmar Herndler-Brandstetter [US], Richard A. Flavell [US], Davor Frleta [US], Cagan Gurer [US], Markus Gabriel Manz [CH], Andrew J. Murphy [US], Noah W. Palm [US], Liang Shan [US], Sean Stevens [US], Till Strowig [DE], George D. Yancopoulos [US], Marcel de Zoete [NL].

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Cytokine Production Assay for TIL

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TIL and tumor target cells were cultured at a 1:1 ratio (1x10⁵ cells each) overnight in round bottom 96-well plates. Autologous tumor cells from enzymatic digestion were used as targets. Supernatants were collected after 24 hours. IFN-gamma was measured using a Human IFNg Quantikine ELISA Kit (R&D Systems, SIF50).

Poch M., Hall M., Joerger A., Kodumudi K., Beatty M., Innamarato P.P., Bunch B.L., Fishman M.N., Zhang J., Sexton W.J., Pow-Sang J.M., Gilbert S.M., Spiess P.E., Dhillon J., Kelley L., Mullinax J., Sarnaik A.A, & Pilon-Thomas S. (2018). Expansion of tumor infiltrating lymphocytes (TIL) from bladder cancer. Oncoimmunology, 7(9), e1476816.

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TIL Characterization and Tumor Reactivity

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Flow cytometric analysis of TIL from each fragment was performed. Expanded TIL was stained with fluorescent antibodies for CD3, CD4, CD8, and CD56 (BD Biosciences, BDB555516). All cells were stained with a Live/Dead Near-IR viability stain (Invitrogen, L10119) and fixed in 2% paraformaldehyde. Data were acquired on an LSR II flow cytometer and analyzed using FlowJo software (TreeStar, Inc.). TIL and autologous tumor cells from enzymatic digestion were cultured at a 1:1 ratio (1 x 10⁵ cells each) overnight in round bottom 96-well plates. Supernatants were collected after 24 h. IFN-gamma was measured using a Human IFNg Quantikine ELISA Kit (R&D Systems, SIF50). Optical density of each well was measured at 450 nm and IFN-γ concentration was calculated from the standard curve. IFN-γ concentration ≥100 pg/ml was the cut-off for reactivity.

Aydin A.M., Bunch B.L., Beatty M., Hajiran A., Dhillon J., Sarnaik A.A., Pilon-Thomas S, & Poch M.A. (2021). The Factors Affecting Expansion of Reactive Tumor Infiltrating Lymphocytes (TIL) From Bladder Cancer and Potential Therapeutic Applications. Frontiers in Immunology, 12, 628063.

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