Immunolabeling of Drosophila Brain Sections

Fly head sections (10 μm thick) were prepared from adults that were frozen in OCT medium (Tissue-Tek, Torrance, CA). Immunolabeling was performed on cryosections sections with mouse anti-24B10 (1:100, DSHB), rat anti-LOVIT (1:100) (Xu and Wang, 2019 (link)), or anti-CSP (1:100, DSHB), rat anti-RFP (1:200, Chromotek, Germany), rabbit anti-Hdc (1:50), rabbit anti-GFP (1:200, Invitrogen, Carlsbad, CA), and rabbit anti-Ebony (1:200, lab of Dr S Carroll, University of Wisconsin, Madison, WI) as primary antibodies. For histamine immunolabeling, the rabbit anti-histamine (1:100, ImmunoStar, Hudson, WI) antibody was pre-adsorbed with carcinine, as previously reported (Xu et al., 2015 (link)). Goat anti-rabbit lgG conjugated to Alexa 488 (1:500, Invitrogen, Carlsbad, CA), goat anti-mouse lgG conjugated to Alexa 488 (1:500, Invitrogen, Carlsbad, CA), goat anti- rabbit lgG conjugated to Alexa 568 (1:500, Invitrogen, Carlsbad, CA), and goat anti-rat lgG conjugated to Alexa 647 (1:500, Invitrogen, Carlsbad, CA) were used as secondary antibodies. The images were recorded with a Zeiss 800 confocal microscope.