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Mouse ifn γ kit

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The BD mouse IFN-γ kit is a laboratory assay used to detect and measure the levels of interferon-gamma (IFN-γ) in mouse samples. IFN-γ is an important cytokine involved in the immune response.

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Lab products found in correlation

Immunospot series 3 analyzer, by Cellular Technology (2 mentions) Pe mhc class 1 pentamer, by ProImmune (2 mentions) Immunospot software, by Cellular Technology (2 mentions) Facsdiva software, by BD (2 mentions) Fixable viability dye efluor 450, by Thermo Fisher Scientific (1 mentions) Lsrii cytometer, by BD (1 mentions) Aec substrate, by Merck Group (1 mentions) Lsrii instrument, by BD (1 mentions) Fitc labeled anti mouse cd8, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

IFN-γ (389 citations) Mouse IFN-γ (7 citations) Ifn-γ ELISPOT mouse kits (2 citations)

4 protocols using mouse ifn γ kit

1

Analyzing Splenocyte Immune Responses

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Splenocytes were analyzed 5 days after the first IT injection of DMXAA or ML RR-S2 CDA. For the ELISPOTs, 106 splenocytes were plated per well and stimulated overnight with SIY peptide (160 nM) or AH1 (1 μM) peptide, with PMA (50 ng/ml) plus ionomycin (0.5 μM) as a positive control, or medium as negative control. Spots were developed using the BD mouse IFN-γ kit according to the manufacturer’s instructions and the number of spots was measured using an Immunospot Series 3 Analyzer and analyzed using ImmunoSpot software (Cellular Technology Ltd). For SIY-pentamer staining, splenocytes were preincubated for 15 min with anti-CD16/32 monoclonal antibody (93) to block potential nonspecific binding, and labeled with PE-MHC class I pentamer (Proimmune) consisting of murine H-2Kb complexed to SIYRYYGL (SIY) peptide, anti–TCRβ-AF700 (H57-597), anti–CD8-Pacific Blue (53-6.7), anti–CD4-Pacific Orange (RM4-5) (all antibodies from BioLegend) and the Fixable Viability Dye eFluor 450 (eBioscience). Stained cells were analyzed using LSR II cytometer with FACSDiva software (BD). Data analysis was conducted with FlowJo software (Tree Star).
Corrales L., Glickman L.H., McWhirter S.M., Kanne D.B., Sivick K.E., Katibah G.E., Woo S.R., Lemmens E., Banda T., Leong J.J., Metchette K., Dubensky TW J.r, & Gajewski T.F. (2015). Direct activation of STING in the tumor microenvironment leads to potent and systemic tumor regression and immunity. Cell reports, 11(7), 1018-1030.
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2

Splenocyte Stimulation and Pentamer Staining

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Splenocytes were plated at 106 cells per well and stimulated overnight with SIY peptide. Spots were developed using the BD mouse IFN-γ kit, and the number of spots was measured using an Immunospot Series 3 Analyzer and analyzed using ImmunoSpot software (Cellular Technology). For pentamer staining, cells were labeled with PE-MHC class I pentamer (Proimmune) consisting of murine H-2Kb complexed to SIYRYYGL (SIY) peptide, anti-CD8-APC (53-6.7), anti-CD19-PerCP-Cy5.5 (6D5), and anti-CD4-PerCP-Cy5.5 (RM4–5). Stained cells were analyzed using FACSCanto or LSR II cytometers with FACSDiva software (BD). Data analysis was conducted with FlowJo software (Tree Star).
Woo S.R., Fuertes M.B., Corrales L., Spranger S., Furdyna M.J., Leung M.Y., Duggan R., Wang Y., Barber G.N., Fitzgerald K.A., Alegre M.L, & Gajewski T.F. (2014). STING-Dependent Cytosolic DNA Sensing Mediates Innate Immune Recognition of Immunogenic Tumors. Immunity, 41(5), 830-842.
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3

Characterizing Tumor-Specific T Cells

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Mice were injected in the subcutaneous flank space with 1×106 tumor cells suspended in 100μl of PBS. Spleens were harvested 7 days after injection for analysis. The enzyme-linked Immunospot assay (ELISPOT) was conducted with the BD mouse IFN-γ kit according to the manufacturer’s protocol. Splenocytes were plated at 106 cells/well and stimulated overnight with, SIINFEKL peptide (160 nM), or PMA (50 ng/ml) and ionomycin (0.5 μM). IFN-γ spots were detected using biotinylated antibody and avidin-peroxidase and developed using AEC substrate (Sigma-Aldrich).
Koss B., Shields B.D., Taylor E.M., Storey A.J., Byrum S.D., Gies A.J., Washam C.L., Choudhury S.R., Ahn J.H., Uryu H., Williams J.B., Krager K.J., Chiang T.C., Mackintosh S.G., Edmondson R.D., Aykin-Burns N., Gajewski T.F., Wang G.G, & Tackett A.J. (2020). Epigenetic control of Cdkn2a.Arf protects tumor-infiltrating lymphocytes from metabolic exhaustion. Cancer research, 80(21), 4707-4719.
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4

Quantification of Antigen-Specific CD8+ T Cells

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1-2 × 106 splenocytes were stained with H-2Kb/SIINFEKL Pro5® Pentamer (ProImmune, UK) for 10 minutes at 22°C. Cells were then washed twice and incubated with FITC-labeled anti-mouse CD8 (eBiosciences) and PECy5-labeled anti-mouse CD19 (ProImmune, UK) antibodies for 20 minutes on ice, shielded from light. Following two further washes, cells were resuspended in fixative and analyzed by flow cytometry using a LSRII instrument (Becton Dickinson, USA). For IFNγ ELISA, 1 × 106 splenocytes were plated and stimulated with SIINFEKEL peptide (10 μg/ml). At 2 days after stimulation, the supernatant was harvested and IFNγ production was estimated by BD mouse Ifnγ kit.
Ahn J., Xia T., Capote A.R., Betancourt D, & Barber G.N. (2018). Extrinsic Phagocyte-Dependent STING-Signaling Dictates the Immunogenicity of Dying Cells. Cancer cell, 33(5), 862-873.e5.
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