Skip to main content
. Author manuscript; available in PMC: 2020 Sep 22.
Published in final edited form as: Sci Transl Med. 2019 Mar 27;11(485):eaau7746. doi: 10.1126/scitranslmed.aau7746

Fig. 5. Specific binding of GPRC5D by scFv clone 109.

Fig. 5.

(A) Binding of HEK293 cells transiently expressing a library of human GPCRs with cytoplasmic GFP to cocultured HEK293 cells transiently expressing anti-GPRC5D scFv clone 109, a long spacer, and cytoplasmic mCherry (both cell types in suspension), quantified by automated flow cytometric analysis. Prespecified threshold for significance (dashed line): Z score = 3, P < 0.0027. (B) Binding of anti-GPRC5D scFv clone 109-mIgG2a Fc chimeric antibody to HEK293 cells expressing the indicated cell surface proteins. Confirmation of binding to potential off-target proteins and nonspecific binders identified in a microarray screen of >4400 transmembrane proteins is shown. ZsGreen1, transfection control; Isotype, irrelevant scFv-mIgG2a Fc, negative control; CTLA-4/CD86 interaction, positive control. (C) Evaluation of GPRC5D(109) CAR activation by potential off-target proteins PCDH1A and FCGR2A. Jurkat Nur77-RFP activation reporter cells expressing a bicistronic plasmid containing a GPRC5D(109) CAR and GFP were cocultured with K562 cells expressing the indicated antigens, GPRC5D (positive control), or BCMA (negative control). Activation is determined as %RFP+GFP+/total GFP+ cells.