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. 2013 Feb 21;121(17):3386–3395. doi: 10.1182/blood-2012-11-468868

Figure 6.

Figure 6

SAP-deficient murine and hu-iNKT cells form fewer conjugates with antigen-pulsed targets. iNKT cells from B6, tamoxifen-treated Sapfl/flCre– or Sapfl/flCre+ mice and EL4 cells were stained with 250 nM carboxyfluorescein succinimidyl ester and bodipy red succinimdyl ester, respectively. iNKT cells were mixed at a 1:1 ratio with PBS44-loaded or unloaded targets, spun down at 1200 rpm for 2 minutes, and incubated at 37°C for varying times. Conjugate frequencies were analyzed by flow cytometry for the presence of cellular aggregates staining positive for bodipy red succinimdyl ester and carboxyfluorescein succinimidyl ester. (A-B) Mean percent iNKT cells (of total iNKT cells) in conjugate with PBS44-loaded or unloaded EL4 cells. (C-D) Mean percent of SAP+ or SAP− iNKT cells ± SEM in conjugate with PBS44-loaded EL4 cells. iNKT cells were flow-sorted from B6 mice (A), tamoxifen-treated Sapfl/flCre− (SAP+) or Sapfl/flCre+ (SAP−) mice (C), or control (SAP+) or SAP-specific (SAP−) siRNA transfected hu-iNKT cells (D). Percent iNKT cells in conjugate was calculated by using the formula [% iNKT-containing conjugates (upper right quadrant)]/[% iNKT-containing conjugates (upper right quadrant) + % iNKT cells not in conjugate (lower right quadrant)] × 100. Data presented are from 3 experiments. Statistical significance was determined by unpaired two-tailed t test. **P < .001.