mouse anti scn2a  (Alomone Labs)

Journal: Nature Communications

Article Title: Proximity analysis of native proteomes reveals phenotypic modifiers in a mouse model of autism and related neurodevelopmental conditions

doi: 10.1038/s41467-024-51037-x

Figure Lengend Snippet: A Schematic illustration of HiUGE-iBioID and its workflow. GS-sgRNA: gene-specific-gRNA. DS-sgRNA: donor-specific-gRNA. Strategies to preserve C-term PDZ-binding motifs of Syngap1, Ctnnb1, Iqsec2, and Lrrc4c are detailed. B Overview of 14 proximity proteomes that segregates according to expected bait functions. C Enrichment analysis of overlapping SFARI genes using hypergeometric probability. The solid red line denotes Bonferroni adjusted p-value at 0.05. D Proximity proteome clustering based on a similarity matrix. E Core proximity proteome between Syngap1 and Anks1b that show highly significant overlaps with SFARI genes. F Core proximity proteome amongst Ank3, Scn2a, and Scn8a that are clustered based on similarity. Modules of proteins were isolated by MCL clustering or GO analysis.

Article Snippet: Antibodies and dilutions consisted of: mouse anti-V5-epitope (ThermoFisher #R960-25, 1:500), rabbit anti-V5-epitope (Cell Signaling #13202 S, 1:1000), guinea pig anti-Homer1 (Synaptic Systems #160004, 1:2000), rabbit anti-Syngap1 (ThermoFisher #PA5-58362, 1:1000), rabbit anti-Shank2 (Cell signaling #12218 S, 1:1000), mouse anti-Ank3 (ThermoFisher #33-8800, 1:1000), mouse anti-Scn2a (Antibodiesinc #75-024, 1:1000), goat anti-mouse Alexa Fluor Plus 594 (ThermoFisher #A32742, 1:1000), goat anti-guinea pig Alexa Fluor 488 (ThermoFisher #A11073, 1:1000), and goat anti-rabbit Alexa Fluor 488 (ThermoFisher #A11008, 1:1000).

Techniques: Binding Assay, Isolation

Journal: Nature Communications

Article Title: Proximity analysis of native proteomes reveals phenotypic modifiers in a mouse model of autism and related neurodevelopmental conditions

doi: 10.1038/s41467-024-51037-x

Figure Lengend Snippet: A Generation of a mouse model based on a clinically identified Scn2a missense mutation (R102Q) in autistic individuals. WES: whole exome sequencing. B Behavioral face-validity of Scn2a +/R102Q mutants was assessed by the zero maze as the percent time and distance traveled in the open areas; the hole-board test as numbers of head pokes and repeated head-pokes; the self-grooming test; and the ultrasonic vocalizations (USVs) as numbers of calls, call durations, and call frequencies during social interaction. No differences were detected in the metrics of pre-social (baseline) responses in the USV tests. * p < 0.05, ** p < 0.01, WT vs . Scn2a +/R102Q mice; independent samples t-tests, two-tailed ( n = 10 or 14 mice per group). Statistics are summarized in Supplementary Data . Plots are mean ± SEM. C Spatial proteomics reveals co-perturbations in Scn2a +/R102Q mutants, two-tailed heteroscedastic t-test on log2-transformed data. MCL analysis discovered a key cluster associated with voltage-gated channel activity that is downregulated, including three targets that intersect with the Scn2a HiUGE-iBioID proximity proteome (underlined). D Scn2a +/R102Q mutant neurons show attenuated activity with the MEA. Scn2a-CRISPRa treatment and a “Combo” treatment with additional expression of SCN1B and FGF12 show differential efficacy in restoring neural activity deficits. * p < 0.05; ** p < 0.01; *** p < 0.001; n.s.: non-significant. One-way ANOVA followed by post-hoc Tukey HSD tests ( n = 48 wells). Plots are mean ± SEM.

Article Snippet: Antibodies and dilutions consisted of: mouse anti-V5-epitope (ThermoFisher #R960-25, 1:500), rabbit anti-V5-epitope (Cell Signaling #13202 S, 1:1000), guinea pig anti-Homer1 (Synaptic Systems #160004, 1:2000), rabbit anti-Syngap1 (ThermoFisher #PA5-58362, 1:1000), rabbit anti-Shank2 (Cell signaling #12218 S, 1:1000), mouse anti-Ank3 (ThermoFisher #33-8800, 1:1000), mouse anti-Scn2a (Antibodiesinc #75-024, 1:1000), goat anti-mouse Alexa Fluor Plus 594 (ThermoFisher #A32742, 1:1000), goat anti-guinea pig Alexa Fluor 488 (ThermoFisher #A11073, 1:1000), and goat anti-rabbit Alexa Fluor 488 (ThermoFisher #A11008, 1:1000).

Techniques: Mutagenesis, Sequencing, Two Tailed Test, Transformation Assay, Activity Assay, Expressing