Immunostaining and Confocal Imaging of MOVAS Cells

We plated MOVAS cells in four-chamber TC slides (Corning) to be used for staining. MOVAS cell staining and imaging was conducted at the Clemson Light Imaging Facility (CLIF), as previously described [29 (link)]. Briefly, cells were incubated with mouse anti-ACTA2 (sc-32251; Santa Cruz Biotechnology) and mouse anti-CD68 (sc-20060; Santa Cruz Biotechnology) primary antibodies, then incubated with Alexa Fluor 546 goat anti-mouse IgG_2a (Invitrogen, Carlsbad, CA, USA) and Alexa Fluor 488 goat anti-mouse IgG₁ (Invitrogen) secondary antibodies. We detected cell nuclei via staining with DAPI (Invitrogen). Cellular imaging was performed using a Leica SP8X MP Confocal System equipped with HyD detectors, a 405 nm laser, a tunable white light laser, and time gating capacity (Leica Microsystems, Buffalo Grove, IL, USA). To capture and export images of the stained MOVAS cells, Leica LAS-X software (Leica Microsystems Version 3.5.5.19976) was utilized.