Ex vivo γδ T cell expansion

γδ T cell were cultured as previously described (28 (link)). Briefly, splenocyte were cultured at 1 × 10⁶ cells per ml in RPMI 1640 containing 10% FCS, antibiotics, 1 X Glutamax (Gibco), 10mM HEPES (Gibco), 1mM sodium pyruvate (Gibco), 55 μM β-mercaptoethanol and non-essential amino acids (Gibco) with 5ng/ml recombinant IL-23 (R&D Systems), 5ng/ml rIL-1β (R&D Systems) and 10μg/ml anti-IFN-g (BioLegend) in 96-well round-bottom plates coated with 1 μg/ml anti-TCR-γδ (clone GL3; Biolegend) for 3 days. Cells were washed and re-seeded on fresh plastic at 1 × 10⁶ cells/ml for a further 3 days as above without TCR-γδ stimulation. Cells were collected at day 6 for flow cytometry analysis.

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Nguyen C.T., Furuya H., Das D., Marusina A.I., Merleev A.A., Ravindran R., Jalali Z., Khan I.H., Maverakis E, & Adamopoulos I.E. (2022). Peripheral γδ T cells regulate neutrophil expansion and recruitment in experimental psoriatic arthritis. Arthritis & rheumatology (Hoboken, N.J.), 74(9), 1524-1534.

Publication 2022

Glutamax HEPES sodium pyruvate recombinant IL-23 rIL-1β anti-IFN-g

Antibiotics Cells Clone Essential amino acids Flow cytometry Hepes Ifn g Mercaptoethanol Ml 23 Pyruvate Sodium Tcr γδ γδ t cell

Corresponding Organization : Harvard University

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Variable analysis

independent variables

Stimulation with anti-TCR-γδ (clone GL3; Biolegend) at 1 μg/ml for 3 days
Culture with 5ng/ml recombinant IL-23 (R&D Systems), 5ng/ml rIL-1β (R&D Systems), and 10μg/ml anti-IFN-g (BioLegend)

dependent variables

Expression of γδ T cell markers (measured by flow cytometry analysis)

control variables

Splenocyte cell culture at 1 × 10^6 cells per ml in RPMI 1640 containing 10% FCS, antibiotics, 1 X Glutamax (Gibco), 10mM HEPES (Gibco), 1mM sodium pyruvate (Gibco), 55 μM β-mercaptoethanol and non-essential amino acids (Gibco)
Cell wash and re-seeding on fresh plastic at 1 × 10^6 cells/ml for a further 3 days without TCR-γδ stimulation

controls

Positive control: Stimulation with anti-TCR-γδ (clone GL3; Biolegend) at 1 μg/ml
Negative control: Not explicitly mentioned

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