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. 2020 Jun 4;8(1):e000266. doi: 10.1136/jitc-2019-000266

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© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.

This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See http://creativecommons.org/licenses/by-nc/4.0/.

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TIGIT but not DNAM1, CD112R or CD96—interacts with Nectin4. (A) FACS staining of IL-2 activated primary NK cells with Nectin4-Ig. Gray filled histogram represents background staining with secondary antibody only; black line histogram represents specific binding as indicated. (B–F) FACS staining of Raji cells transfected either with an empty vector as control (gray histograms) or with Nectin4 (black histograms). Cells were stained with commercial anti-Nectin4 mAb (B), TIGIT-Ig (C), DNAM1-Ig (D), CD112-Ig (E) or CD96-Ig (F). Figures show one representative experiment out of three performed. Graph depicting the mean fluorescence intensity values of the stainings appears in online supplementary figure 2. (G) Direct binding of Nectin4 to TIGIT. The binding of fluorophore-labeled TIGIT-Ig and its ligands PVR-Ig (red) and Nectin4-Ig (green) was determined using MST. Measurements were repeated with at least three independent protein preparations. FACS, fluorescence-activated cell sorting; IL-2, interleukin-2; MST, microscale thermophoresis; NK, natural killer.