ELISpot assays were performed following manufacturer’s protocols using the Mouse IFN-γ Enzyme-Linked ImmunoSpot (ELISpot) set (BD Biosciences Cat. No. 551083). In brief, serial dilutions of lymphoid cells from the spleen, draining lymph node, or TILs were cultured in RPMI-CM with or without AH1 tumor antigen peptide (2 µg/mL; ChiScientific, sequence: SPSYVYHQF) and 10-fold lower numbers of BMDCs in anti-IFN-γ-coated (5 µg/mL antibody incubation overnight, BD Biosciences Cat. No. 551881) 96-well ELISpot plates (Millipore Sigma Cat. No. S2EM004M99) for 19–24 hours at 37°C in 5% CO2. Secretion of IFN-γ was detected using biotinylated anti-IFN-γ (2 µg/mL, BD Biosciences Cat. No. 551881) followed by the addition of Streptavidin-HRP (1:100, BD Biosciences Cat. No. 557630) and AEC Chromogen/Substrate (BD Biosciences, Cat. No. 551951). ELISpots were visualized using ImmunoSpot S6 Universal and quantified using ImmunoSpot V.7.0.30.4 Analyzer Professional DC software.
Mouse ifn γ enzyme linked immunospot elispot set
Manufactured by BD
The Mouse IFN-γ Enzyme-Linked ImmunoSpot (ELISpot) set is a laboratory equipment product that enables the detection and quantification of mouse interferon-gamma (IFN-γ) secreting cells in a sample. The set provides the necessary reagents to perform the ELISpot assay, a widely used technique for the analysis of cellular immune responses.
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Lab products found in correlation
Anti ifn γ, by BD (1 mentions)
Aec chromogen substrate, by BD (1 mentions)
Streptavidin hrp, by BD (1 mentions)
Ctl immunospot s5 versa analyzer, by Cellular Technology (1 mentions)
Aec substrate set, by BD (1 mentions)
Mouse ifn γ elispot set, by BD (1 mentions)
Dimethyl sulfoxide (dmso), by Nacalai Tesque (1 mentions)
2 protocols using mouse ifn γ enzyme linked immunospot elispot set
1
In Vitro Antigen-Specific T Cell Assay
2
Enhancing WT1-Specific T Cell Immunity
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To assess the ability of adegramotide to enhance nelatimotide-mediated induction of WT1-specific cytotoxic T cells, 8-week-old female HLA-A*02:01 transgenic mice received 2 weekly intradermal injections of either DSP-7888 Emulsion or nelatimotide-only Emulsion. The dose of nelatimotide in DSP-7888 Emulsion and nelatimotide-only emulsion was 1 mg/head. The dose of adegramotide in DSP-7888 Emulsion was 0.75 mg/head. Mice were euthanized by carbon dioxide inhalation 7 days after the second injection. Splenocytes from each mouse were transferred in triplicate to 96-well plates coated with anti-mouse IFN-γ, as provided in the Mouse IFN-γ Enzyme-Linked Immunospot (ELISPOT) Set (BD Biosciences), and seeded at 1.25 × 105 cells/well. Cells were then stimulated with 10 μg/mL WT1126–134 peptide (Life Technologies) or 0.025% volume DMSO (Nacalai Tesque, Inc.) and incubated for 19 h. IFN-γ was detected by biotinylated anti-mouse IFN-γ and streptavidin-HRP from the Mouse IFN-γ ELISPOT Set and AEC Substrate Set (BD Biosciences). The number of spots in each well was determined using CTL-ImmunoSpot S5 Versa Analyzer (Cellular Technology, Ltd.) and quantified using the Smart Count mode in software version 5.0.3. T-cell induction in the two treatment groups was compared via two-sided t test using SAS version 9.4.
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