Sc 166274

The following antibodies were used: anti-PAD2 antibody (PA5-19474, Thermo Scientific; diluted 1 : 100) and anti-PAD4 (PA5-2217, Thermo Scientific; diluted 1 : 100), anti-Ro60 (H-300 Sc-20961) or anti-La (sc-166274, Santa Cruz Biotechnology; diluted 1 : 100), and anti-β-actin (ab2072 Abcam). Peroxidase-conjugated goat anti-mouse IgG (Sigma, St. Louis, MO) or goat anti-rabbit IgG (Abcam ab759) was used as secondary antibody. Additionally, an anti-citrulline antibody (231246, Calbiochem, Darmstadt, Germany; 1 : 100 dilution in 10% foetal bovine serum (FBS) PBS) was used for immunohistochemistry.

Exosomes were adsorbed on the nickel grids by incubating at 37 °C for 30 min. The grids were washed in 100 µl drop of 1X PBS for 5 min and then transferred into a 100 µl drop of 0.5 M NH₄CL for 3 min. Blocking was done by placing the grid in 5% BSA for 20 min. Grids were washed and incubated overnight at 4 °C in anti-CD63 monoclonal antibody (ab8219, Abcam, UK) or anti-La monoclonal antibody (sc-166274, Santa Cruz, USA) or anti-EBNA1 monoclonal antibody (MA1-7271, Thermofisher, USA) (negative control). Each antibody was used at 1:25 dilution. Next day, the grids were washed and then incubated in secondary antibody (anti-mouse IgG conjugated to 10-nm gold particles, TAAB, UK) at 1:100 dilution for 2 hours. The grids were washed and fixed in 2.5% aqueous glutaraldehyde for 5 min, then contrasted with 2% uranyl acetate for 5 min and examined using Philips CM10 transmission electron microscope (TEM) (Philips, the Netherland) at 80 K volts.