Rabbit anti vdac

Microglia (1 × 10⁵) were plated on poly-l-lysine glass coverslips (Fisher Scientific, Pittsburgh, PA, 08-774-383) in 12-well plates. Cells were treated for 18 h with vehicle (media) or 100 ng/mL LPS in media supplemented with 2% FBS (2 mL solution per well; Hyclone, GE Healthcare Life Sciences, Marlborough, MA). Immunocytochemistry was performed via conventional techniques. That is, cells were fixed with 4% paraformaldehyde (PFA) and permeabilized with 0.2% Triton X-100. Blocking was performed with 10% normal donkey serum (Jackson Laboratories, Bar Harbor, ME) for 3 h before being incubated with primary antibodies. Goat anti-TSPO (Abcam, Cambridge, MA, ab118913, RRID:AB_10898989, 1:500); mouse anti-gp91^phox (BD Biosciences, San Jose, CA, 611415, RRID:AB_398937, 1:100); mouse anti-p22^phox (Santa Cruz Biotechnology, Dallas, TX, sc-130,551, RRID:AB_2245805, 1:100); rabbit anti-VDAC (Abcam, Cambridge, MA, ab15895, RRID:AB_2214787, 1:500) were used. After washing, cells were incubated with appropriate Alexa Fluor secondary antibodies for 1 h (Life Technologies, Carlsbad, CA; AF 488, 594, and 647, 1:500). Coverslips were mounted with Prolong with DAPI (Life Technologies, Carlsbad, CA) to counterstain for cell nuclei.

Rat anti-HA (100 µg/ml, clone 3F10, Roche), mouse anti-HA (1 mg/ml, clone HA.11, 16B12 Covance), rabbit anti-IFITM1-NTD (100 µg/ml, Sigma), rabbit anti-IFITM3-NTD (250 µg/ml, Abgent), rabbit anti-VDAC (1 mg/ml, Abcam), rabbit anti-calreticulin (Thermo Scientific), mouse anti-tubulin (clone DM1A, ascites fluid, Sigma), goat anti-rat Alexa-488, goat anti-rabbit Alexa-488, goat anti-mouse Alexa-594 and goat anti-rabbit Alexa-647 (all 2 mg/ml, Life Technologies), goat anti-rabbit IRDye 680 and goat anti-mouse IRDye 800 (1 mg/ml, Li-COR), and wheat germ agglutinin (WGA) conjugated to Alexa-647 fluorophore (1 mg/ml, Invitrogen), were used at the dilutions given below.