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  • 99
    Thermo Fisher phusion hot start dna polymerase
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    Millipore α bungarotoxin fitc
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Roche expand long range dntpack
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    NEN Life Science biotinylated tyramide
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Absolute Antibody Ltd anti tissue factor 5g9
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    BioFire Defense filmarray ngds bt e assay
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Jackson Immuno alexa 488 conjugated strepavidin
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Thermo Fisher electrophoretic mobility shift assay
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Santa Cruz Biotechnology anti ck10
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Millipore h2 o2
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    baseclick GmbH edu hts kit
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Roche lightmix ebola zaire rrt pcr test
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Altona Diagnostics realstar filovirus screen rt pcr kit
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Millipore palmitoyl l carnitine
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Becton Dickinson intracellular cytokine analysis
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Vector Laboratories pbs
    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with <t>α-bungarotoxin-FITC</t> (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.
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    Thermo Fisher α bungarotoxin
    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    Millipore lipopolysaccharide
    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    Becton Dickinson facscalibur
    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    Dojindo Labs annexin v fitc apoptosis kit
    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    Becton Dickinson cell fixation permeabilization kits
    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    Millipore calcium ionomycin
    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and <t>α-bungarotoxin</t> staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p
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    Image Search Results


    Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with α-bungarotoxin-FITC (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.

    Journal: PLoS ONE

    Article Title: Effect of Delayed Peripheral Nerve Repair on Nerve Regeneration, Schwann Cell Function and Target Muscle Recovery

    doi: 10.1371/journal.pone.0056484

    Figure Lengend Snippet: Neuromuscular junctions and expression of nicotinic acetylcholine receptors (nAChRs). (A) Muscle sections were stained for nAChRs with α-bungarotoxin-FITC (green) and with SV2A antibody (red) to mark pre-synaptic structures. Note the coincident staining of both markers indicating presumptive functional NMJs. DAPI staining (blue) shows nuclei. Scale bar = 25 µm. (B) RT-PCR analysis of the different nAChR subunits (α, β, γ, δ, ε) and muscle specific kinase MuSK in control muscle (con) and muscle from the operated side of animals undergoing immediate (0) or 1,3,6 months delayed nerve repair. 18 S was used as a house-keeping gene.

    Article Snippet: Following washing, the slides were then incubated for 1 h in the dark with secondary antibody Alexa Fluor 568 goat anti-rabbit IgG (1∶100, Invitrogen) and α-bungarotoxin-FITC (1∶10, Sigma).

    Techniques: Expressing, Staining, Functional Assay, Reverse Transcription Polymerase Chain Reaction

    The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and α-bungarotoxin staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p

    Journal: Scientific Reports

    Article Title: R-spondin 2 promotes acetylcholine receptor clustering at the neuromuscular junction via Lgr5

    doi: 10.1038/srep28512

    Figure Lengend Snippet: The Rspo2/Lgr5 complex induces activation and phosphorylation of MuSK and AChR clustering. ( A ) Lgr5 immunostaining and α-bungarotoxin staining of the NMJs as in Fig. 2B . ( B,C ) Lgr5 was co-immunoprecipitated with anti-myc (B) or anti-Flag (C) antibody in HEK293 cells. The normalized ratio of co-immunoprecipitated Lgr5 to total Lgr5 is shown in Supplementary Fig. S3A and S3B , and the mean value is indicated below the blot. ( D ) ATF2-luciferase reporter assays as in Fig. 2D in siRNA-transfected HEK293 cells. RLA are normalized to that of BSA with siControl. Mean and SD are indicated (** p

    Article Snippet: After the removal of the connective tissue, the whole-mount diaphragm was permeabilized with 0.5% Triton X-100 in PBS for 10 min and then incubated overnight with α-bungarotoxin conjugated with the Biotin-XX Microscale Protein Labeling Kit (1:800, Invitrogen), anti-peripherin antibody (1:800, Millipore, AB1530), and anti-synaptophysin antibody (1:100, Invitrogen, 180130).

    Techniques: Activation Assay, Immunostaining, Staining, Immunoprecipitation, Luciferase, Transfection

    Rspo2 is enriched at the NMJ and activates MuSK to induce AChR clustering. ( A ) Rspo2 expression in the diaphragm and spinal cord normalized by Gapdh and also to E18.5 diaphragm. Mean and SD ( n = 3) are indicated. ( B ) Rspo2 immunostaining and α-bungarotoxin staining for AChR at the NMJ of the tibialis anterior muscle (cross section). ( C ) AChR clusters visualized with α-bungarotoxin (red). C2C12 myotubes were cultured with 0.05 nM agrin and/or 0.05 nM Rspo2. Arrows point to the AChR clusters with an axis length of 4 μm or more. Blinded morphometric analysis is shown in Supplementary Fig. S2A . ( D ) ATF2-luciferase reporter assay to quantify agrin (10 ng/ml)- and Rspo2 (100 ng/ml)-mediated activation of MuSK signaling in transfected HEK293 cells. Relative luciferase activities (RLA) are normalized to that with empty vectors. Mean and SD are indicated ( n = 3). ** p

    Journal: Scientific Reports

    Article Title: R-spondin 2 promotes acetylcholine receptor clustering at the neuromuscular junction via Lgr5

    doi: 10.1038/srep28512

    Figure Lengend Snippet: Rspo2 is enriched at the NMJ and activates MuSK to induce AChR clustering. ( A ) Rspo2 expression in the diaphragm and spinal cord normalized by Gapdh and also to E18.5 diaphragm. Mean and SD ( n = 3) are indicated. ( B ) Rspo2 immunostaining and α-bungarotoxin staining for AChR at the NMJ of the tibialis anterior muscle (cross section). ( C ) AChR clusters visualized with α-bungarotoxin (red). C2C12 myotubes were cultured with 0.05 nM agrin and/or 0.05 nM Rspo2. Arrows point to the AChR clusters with an axis length of 4 μm or more. Blinded morphometric analysis is shown in Supplementary Fig. S2A . ( D ) ATF2-luciferase reporter assay to quantify agrin (10 ng/ml)- and Rspo2 (100 ng/ml)-mediated activation of MuSK signaling in transfected HEK293 cells. Relative luciferase activities (RLA) are normalized to that with empty vectors. Mean and SD are indicated ( n = 3). ** p

    Article Snippet: After the removal of the connective tissue, the whole-mount diaphragm was permeabilized with 0.5% Triton X-100 in PBS for 10 min and then incubated overnight with α-bungarotoxin conjugated with the Biotin-XX Microscale Protein Labeling Kit (1:800, Invitrogen), anti-peripherin antibody (1:800, Millipore, AB1530), and anti-synaptophysin antibody (1:100, Invitrogen, 180130).

    Techniques: Expressing, Immunostaining, Staining, Cell Culture, Luciferase, Reporter Assay, Activation Assay, Transfection

    Apposition of nerve terminals and muscle endplates is compromised in the E18.5 diaphragm of R-spondin 2 ( Rspo2) −/− mice. ( A–F ) Representative confocal images of the left diaphragm at E18.5 labeled with an anti-synaptophysin antibody and α-bungarotoxin to visualize the nerve terminals and acetylcholine receptors (AChRs), respectively. Endplates of the wild-type muscles were mostly ovoid-shaped ( B ), whereas the endplates of Rspo2 −/− muscles were large, round, and heterogeneously stained ( E ). ( G–I ) Blinded morphometric analysis of synaptophysin ( G ) and AChR ( H ) in the AChR clusters revealed that NMJ areas were markedly enlarged at E18.5. Numbers of synaptophysin-positive clusters ( G ) and AChR-positive clusters ( H ) are shown. ( I ) The ratio is calculated by dividing the synaptophysin-positive area by the AChR-positive area. Note that not all AChR-positive (red) pixels were synaptophysin-positive (green) in each AChR cluster. Mean and standard deviation (SD; n = 6) are indicated. **** p

    Journal: Scientific Reports

    Article Title: R-spondin 2 promotes acetylcholine receptor clustering at the neuromuscular junction via Lgr5

    doi: 10.1038/srep28512

    Figure Lengend Snippet: Apposition of nerve terminals and muscle endplates is compromised in the E18.5 diaphragm of R-spondin 2 ( Rspo2) −/− mice. ( A–F ) Representative confocal images of the left diaphragm at E18.5 labeled with an anti-synaptophysin antibody and α-bungarotoxin to visualize the nerve terminals and acetylcholine receptors (AChRs), respectively. Endplates of the wild-type muscles were mostly ovoid-shaped ( B ), whereas the endplates of Rspo2 −/− muscles were large, round, and heterogeneously stained ( E ). ( G–I ) Blinded morphometric analysis of synaptophysin ( G ) and AChR ( H ) in the AChR clusters revealed that NMJ areas were markedly enlarged at E18.5. Numbers of synaptophysin-positive clusters ( G ) and AChR-positive clusters ( H ) are shown. ( I ) The ratio is calculated by dividing the synaptophysin-positive area by the AChR-positive area. Note that not all AChR-positive (red) pixels were synaptophysin-positive (green) in each AChR cluster. Mean and standard deviation (SD; n = 6) are indicated. **** p

    Article Snippet: After the removal of the connective tissue, the whole-mount diaphragm was permeabilized with 0.5% Triton X-100 in PBS for 10 min and then incubated overnight with α-bungarotoxin conjugated with the Biotin-XX Microscale Protein Labeling Kit (1:800, Invitrogen), anti-peripherin antibody (1:800, Millipore, AB1530), and anti-synaptophysin antibody (1:100, Invitrogen, 180130).

    Techniques: Mouse Assay, Labeling, Staining, Standard Deviation

    Lack of R-spondin 2 (Rspo2) in mice has minimal effects on spinal motor neuron (SMN) survival and muscle differentiation, but has a significant effect on acetylcholine receptor (AChR) clusters in the left diaphragm. ( A ) Immunostaining for islet1/2 expressed in the SMNs of the spinal cord (C3-C6) at embryonic day ( E ) 18.5. ( B ) The number of islet1/2-positive SMNs in wild-type (+/+), heterozygous Rspo2 -knockout (+/−), and homozygous Rspo2 -knockout (−/−) mice. Bars indicate the mean and standard error of mean (SE) ( n > 90). No statistically significant differences (n.s.) were observed with one-way ANOVA. ( C,D ) Hematoxylin and eosin staining of the tibialis anterior muscle of mice at E18.5. ( E–H ) Representative electron micrographs of the diaphragms at E18.5 at different magnifications. ( I ) Thickness of muscle fibers at the Z disk in the left diaphragms of wild-type (+/+) and Rspo2 -knockout (−/−) mice. Five to seven electron micrographs were analyzed in each mouse. ( J ) Surface views of the left diaphragms harvested from wild-type (+/+) and Rspo2 -knockout (−/−) mice at E18.5. AChR was stained with Alexa546-conjugated α-bungarotoxin (red). The widths of the AChR bands of wild-type and Rspo2 -kockout diaphragms were 143.34 ± 3.73 μm and 221.85 ± 6.52 μm, respectively ( p

    Journal: Scientific Reports

    Article Title: R-spondin 2 promotes acetylcholine receptor clustering at the neuromuscular junction via Lgr5

    doi: 10.1038/srep28512

    Figure Lengend Snippet: Lack of R-spondin 2 (Rspo2) in mice has minimal effects on spinal motor neuron (SMN) survival and muscle differentiation, but has a significant effect on acetylcholine receptor (AChR) clusters in the left diaphragm. ( A ) Immunostaining for islet1/2 expressed in the SMNs of the spinal cord (C3-C6) at embryonic day ( E ) 18.5. ( B ) The number of islet1/2-positive SMNs in wild-type (+/+), heterozygous Rspo2 -knockout (+/−), and homozygous Rspo2 -knockout (−/−) mice. Bars indicate the mean and standard error of mean (SE) ( n > 90). No statistically significant differences (n.s.) were observed with one-way ANOVA. ( C,D ) Hematoxylin and eosin staining of the tibialis anterior muscle of mice at E18.5. ( E–H ) Representative electron micrographs of the diaphragms at E18.5 at different magnifications. ( I ) Thickness of muscle fibers at the Z disk in the left diaphragms of wild-type (+/+) and Rspo2 -knockout (−/−) mice. Five to seven electron micrographs were analyzed in each mouse. ( J ) Surface views of the left diaphragms harvested from wild-type (+/+) and Rspo2 -knockout (−/−) mice at E18.5. AChR was stained with Alexa546-conjugated α-bungarotoxin (red). The widths of the AChR bands of wild-type and Rspo2 -kockout diaphragms were 143.34 ± 3.73 μm and 221.85 ± 6.52 μm, respectively ( p

    Article Snippet: After the removal of the connective tissue, the whole-mount diaphragm was permeabilized with 0.5% Triton X-100 in PBS for 10 min and then incubated overnight with α-bungarotoxin conjugated with the Biotin-XX Microscale Protein Labeling Kit (1:800, Invitrogen), anti-peripherin antibody (1:800, Millipore, AB1530), and anti-synaptophysin antibody (1:100, Invitrogen, 180130).

    Techniques: Mouse Assay, Immunostaining, Knock-Out, Staining