Proliferation was quantified with a CFSE dilution assay. T cells were washed and stained with 1 μmol/L CFSE (Life Technologies) for 3.5 minutes at room temperature. The reaction was quenched by adding PBS with 10% FBS and washed twice with same buffer. Cells were then incubated at a ratio of 1:1 with irradiated target cells (K562-CD19 vs. K562-wild type) for 120 hours in a cytokine-free media. Live CD3+ cells were then enumerated using bead-based flow cytometry as previously described (32 (link)). The fold change of live T cells was calculated relative to the live T-cell count at day 0. Supernatants were also collected at 24 hours to assess cytokine production. Measurement of Cytokine was performed with a Luminex bead array platform (Life Technologies) according to the manufacturer’s instructions. All samples were analyzed in triplicate and compared against multiple internal standards with a 9-point standard curve. Data were acquired on a FlexMAP-3D system (Luminex), and analysis was performed in XPonent 4.0 software (Luminex), as well as through five-parameter logistic regression (4 (link)).
Luminex bead array platform
Manufactured by Thermo Fisher Scientific
The Luminex bead array platform is a multiplex assay system that allows for the simultaneous detection and quantification of multiple analytes in a single sample. The platform utilizes color-coded magnetic beads coated with specific capture agents to bind and detect target molecules. This technology enables efficient and high-throughput analysis of a wide range of biomolecules, including proteins, nucleic acids, and other analytes.
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Lab products found in correlation
5 protocols using luminex bead array platform
1
T Cell Proliferation Quantification
2
Cytokine Production Assay with T Cells
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T cells were incubated with irradiated target cells at a ratio of 1:1 (1 × 106 cells per ml each) in cytokine-free medium. Supernatants were collected after 24 h to assess the cytokine production. The cytokine measurements were performed using a Luminex bead array platform (Life Technologies) according to the manufacturer’s instructions39 (link).
3
Luminex-based Cytokine Profiling
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HS versus Phx cytokine levels were analyzed with a Luminex bead array platform (Life Technologies) according to the manufacturer’s instructions. All samples were analyzed in triplicate and compared against multiple internal standards with a nine-point standard curve. Data were acquired on a FlexMAP-3D system (Luminex), and analysis was performed in XPonent 4.0 software (Luminex), as well as through five-parameter logistic regression.32 (link)
4
Quantifying Secreted IFNγ by ELISA
5
Quantifying Secreted IFNγ by ELISA
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Secreted IFNγ in coculture supernatants was detected by ELISA (BioLegend) or with a Luminex bead array platform (Thermo Fisher) according to the manufacturer’s instructions. All samples were analyzed in triplicate and compared against multiple internal standards, with a seven-point standard curve.
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