Quantikine elisa kit for murine cxcl10

Tumour cells were seeded overnight on a 96-well plate (2 × 10⁴ cells per well in RPMI). Recombinant mouse interferon-γ (R&D Systems) was added at either 1000 or 5000 Units ml⁻¹ for 24 or 48 h and media were collected. Samples were assayed in triplicate using a Quantikine ELISA kit for murine CXCL10 (R&D Systems).

Stable CXCL10 gene suppression was performed using TRC Lentiviral shRNA clones TRCN0000068210 (sequence: 5′-TAGATTCCGGATTCAGACATC-3′ referred as CXCL10 KD #1) and TRCN0000068212 (sequence: 5′-TTGATGGTCTTAGATTCCGGA-3′ referred as CXCL10 KD #2) (Dharmacon, Pittsburgh, PA, USA). Stable CXCR3 gene suppression was achieved using TRC Lentiviral shRNA clones TRCN0000027391 (sequence: 5′-TTCTCTCCGTGAAGATGACGG-3′ referred as CXCR3 KD #1) and TRCN0000027383 (sequence: 5′-TTTCTCGACCACAGTTGCGGG-3′ referred as CXCR3 KD #2). Scrambled shRNAs served as nontargeting controls. Suppression was confirmed using western blot analysis and the Quantikine ELISA kit for murine CXCL10 (R&D Systems).