Fluorescent Labeling of C2-Crry and Anti-CII Antibodies

To show the binding of C2-Crry in vitro to apoptotic FLS, we labeled the protein with IRDye800-NHS ester (Li-COR Biosciences) as we have shown previously (60; 61 (link)). C2-Crry fusion protein was incubated with a 10-fold molar excess of dye for 5 h at 4°C in PBS. Unbound dye was removed with a 7 kDa Zeba desalting column (Thermo Fisher). After labeling C2-Crry with IRDye800 and addition to a Zeba column, the protein was eluted from the column with PBS and stored at 4°C before use according to our previously published study (60; 61 (link)). Similarly, Arthrogen (anti-CII) mAbs were labeled with IRDye 800 (60; 61 (link)).