Chromium release assay

NK cell effector cells were co-cultured with target cells that had been pre-incubated with 100 µCi ⁵¹Cr for 1 or 4 hr in 96-well round-bottomed plates at 37°C 5% CO₂. 1% IGEPAL (v/v) (Sigma-Aldrich) was used to lyse maximal release control wells and plates were centrifuged. Supernatant was transferred to a LUMA plate (Perkin Elmer) and dried overnight. Plates were read with a TopCount NXT and % specific lysis was calculated as follows: (sample – average spontaneous release) / (average total release – average spontaneous release) x 100. For experiments done with Pyk2 inhibition, effectors were pre-incubated for 10 min with 5 µM PF431396 (Tocris) then assays were performed in the presence of 5 µM PF431396 or equivalent volume of DMSO as a vehicle control.