Immunocytochemistry of SH-SY5Y and HEK293T Cells

Immunocytochemistry (ICC) was performed as described in Perland et al. (2016) (link) on transfected SH-SY5Y cells and wild-type HEK293T cells (only anti-UNC93A) with antibodies all diluted in anti-UNC93A [rabbit, 1:100ab69443, Abcam, (Ceder et al., 2017 (link))], anti-KCNK3 [mouse, 1:100, ab186352 Abcam, (Schmidt et al., 2015 (link))], anti-KCNK9 (mouse, 1:100, Sigma, K0514), and anti-Flag [mouse, 1:200, F3165, Sigma, (Sriramachandran et al., 2019 )]. Images were acquired using Olympus microscope BX55 with an Olympus DP73 camera and the cellSens Dimension v1.14 (Olympus) and were then handled using ImageJ, Fiji edition (Schindelin et al., 2012 (link)). The epitope against UNC93A was pair-wise aligned against the three isoforms of CG4928 (FBpp0074118, FBpp0074119, and FBpp0309738) using EMBOSS Needle (global alignment) and EMBOSS Water (local alignment) to investigate protein identity and similarity (Li et al., 2015 (link)) (Supplementary Dataset S4).