Journal: The Journal of Neuroscience

Article Title: Identification of VAPA and VAPB as Kv2 Channel-Interacting Proteins Defining Endoplasmic Reticulum–Plasma Membrane Junctions in Mammalian Brain Neurons

doi: 10.1523/JNEUROSCI.0893-18.2018

Figure Lengend Snippet: Antibody information

Article Snippet: N106/36 , Full-length recombinant human ankG , Mouse IgG2b mAb, NeuroMab catalog #75-147, RRID: AB_10675130 , Tissue culture supernatant, 1:5 , 9 9.

Techniques: Concentration Assay, Affinity Purification, Purification, Recombinant

Journal: The Journal of Neuroscience

Article Title: Identification of VAPA and VAPB as Kv2 Channel-Interacting Proteins Defining Endoplasmic Reticulum–Plasma Membrane Junctions in Mammalian Brain Neurons

doi: 10.1523/JNEUROSCI.0893-18.2018

Figure Lengend Snippet: CRISPR-mediated KO of VAPA expression affects Kv2.1 localization in RAW264.7 cells A. Representative TIRF images of SEP-Kv2.1 expression in a live WT RAW264.7 cell expressing SEP-Kv2.1 (green is cell surface pHluorin fluorescence, magenta is total cellular mCherry fluorescence). Scale bar is 5 μm and holds for all panels in figure. B, Representative TIRF images of GFP-Kv2.1 (green) and BFP-SEC61β (magenta) in live WT (left) or VAPA KO (right) RAW264.7 cells coexpressing GFP-Kv2.1 and BFP-SEC61β. Note the reduction in clustered Kv2.1 expression in the VAPA KO cell. C, Line scan analysis of selection indicated in merged image of WT RAW264.7 cell coexpressing GFP-Kv2.1 and BFP-SEC61β. D, Line scan analysis of selection indicated in merged image of VAPA KO RAW264.7 cell coexpressing GFP-Kv2.1 and BFP-SEC61β. E, Representative TIRF image of live WT (left) or VAPA KO (right) RAW264.7 cells expressing GFP-Kv1.4. F, Summary graph of CV measurements of GFP-Kv2.1 in WT and VAPA KO RAW264.7 cells (****p = 6.648 × 10−9, n = 29 cells, two-tailed unpaired t test). G, Summary graph of CV measurement of GFP-Kv1.4 in WT and VAPA KO RAW264.7 cells (ns, 0.5603, n = 10 cells, two-tailed unpaired t test). H-K. Images of WT (left) or VAPA KO (right) RAW264.7 cells used for high-content analysis of immunolabeling (magenta) and with Hoechst labeling of nuclei (green). H, No primary (anti-IgG2b secondary antibody). I, Positive control anti-mortalin mAb N52A/42 (IgG1). J, Anti-VAPA mAb N479/22 (IgG2a). K, Anti-VAPA/B mAb N479/107 (IgG2b). Scale bar in H is 65 μm and holds for H–K. L, Mean immunolabeling intensity of all samples across 1300–2800 individual cells in each sample. See Materials and Methods for further details of these antibody validation results.

Article Snippet: N106/36 , Full-length recombinant human ankG , Mouse IgG2b mAb, NeuroMab catalog #75-147, RRID: AB_10675130 , Tissue culture supernatant, 1:5 , 9 9.

Techniques: CRISPR, Expressing, Fluorescence, Selection, Two Tailed Test, High Content Screening, Immunolabeling, Labeling, Positive Control, Biomarker Discovery