ciliary neurotrophic factor (Alomone Labs)

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Alomone Labs ciliary neurotrophic factor

Syt1 is found in close proximity to the ciliary <t>neurotrophic</t> factor receptor (Cntfr) and regulates synaptic vesicle dynamics. a-c ) NbLumSyt1-APEX2 allows efficient biotinylation of proteins in the proximity of Syt1 upon live uptake in hippocampal neurons to facilitate live-cell proteomic mapping. Representative images of neurons upon uptake of NbLumSyt1-APEX2, where biotinylated proteins are revealed with fluorescent streptavidin (a) . In the absence of H 2 O 2 , only a few endogenous biotinylated proteins are observable. In the presence of all the components, the reaction occurred efficiently, as revealed in western blot analysis of labeled neurons ( b ). To identify the interactors of Syt1, in situ proximity labeling was performed with NbLumSyt1-APEX2 ( c ). The electron microscopy image in the scheme is an example of the labeled vesicles, as revealed upon photoconverting 3,3’-diaminobenzidine (DAB) into a stable, electron microscopically visible dark product. d ) Protein intensities measured with LC‒MS/MS at the input and upon enrichment of the biotinylated proteins. Two controls were used: neurons without nanobodies or neurons where an unrelated nanobody (anti-ALFA-Nb) was provided in the medium. Note that since the primary neurons do not express the ALFA tag, this control will reveal the effect of the unspecific biotinylation of the membranes occurring during the labeling period. Note that Syt1, as expected, is efficiently biotinylated and enriched upon IP with streptavidin beads. See methods for details concerning the experiments and analyses. e ) Summary of the gene ontologies (GOs; cellular components) for the proteins biotinylated upon live uptake of NbLumSyt1-APEX2 (for a detailed list, see Supplementary Table 1). As expected, synaptic components and membrane GO terms were overrepresented. f ) Possible interactors identified via live-cell proteomic mapping and enrichment vs. input and vs. IP control. Cntfr was found to be the most enriched candidate, together with other proteins that could be studied in future works. g ) Super-resolution stimulation emission depletion (STED) imaging reveals that ~ 20% of boutons labeled with live uptake are also positive for Cntfr. In this case, for cross-validation purposes, live uptake was performed with the 604.2 Syt1-luminal antibody. h ) Proximity ligation assay (in situ PLA) using antibodies against the luminal portion of Syt1 and anti-Cntfr confirmed the close proximity of these two proteins. A primary antibody against a protein not expressed in hippocampal neurons (Ribeye) was used as the negative control. i ) Blocking the network activity of primary hippocampal neurons with tetrodotoxin (TTX) for 2 h decreases the in situ PLA signal between Syt1 and Cntfr. Stimulation with the ligand of Cntfr <t>(Cntf;</t> 8 nM) for 2 h does not change the PLA signal between Syt1 and Cntfr. j , k ) Stimulation of neurons with Cntf for 24 h increases SV exo-endocytosis. Scale bars: 10 μm in a ; 500 nm in c ; 5 μm in g-j . The error bars indicate the means ± SEMs for panel d, and the 5th or 95th percentile for box plots; ** p < 0.01; *** p < 0.001

Ciliary Neurotrophic Factor, supplied by Alomone Labs, used in various techniques. Bioz Stars score: 94/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/ciliary+neurotrophic+factor+cntf/Recombinant+rat+CNTF+protein/pmc13185310-442-24-29
Average 94 stars, based on 11 article reviews

ciliary neurotrophic factor - by Bioz Stars, 2026-09

94/100 stars

Images

1) Product Images from "A versatile nanobody platform for live and super-resolution imaging of synaptic vesicle dynamics and plasticity in rodent and human neurons"

Article Title: A versatile nanobody platform for live and super-resolution imaging of synaptic vesicle dynamics and plasticity in rodent and human neurons

Journal: Journal of Nanobiotechnology

doi: 10.1186/s12951-026-04489-w

Syt1 is found in close proximity to the ciliary neurotrophic factor receptor (Cntfr) and regulates synaptic vesicle dynamics. a-c ) NbLumSyt1-APEX2 allows efficient biotinylation of proteins in the proximity of Syt1 upon live uptake in hippocampal neurons to facilitate live-cell proteomic mapping. Representative images of neurons upon uptake of NbLumSyt1-APEX2, where biotinylated proteins are revealed with fluorescent streptavidin (a) . In the absence of H 2 O 2 , only a few endogenous biotinylated proteins are observable. In the presence of all the components, the reaction occurred efficiently, as revealed in western blot analysis of labeled neurons ( b ). To identify the interactors of Syt1, in situ proximity labeling was performed with NbLumSyt1-APEX2 ( c ). The electron microscopy image in the scheme is an example of the labeled vesicles, as revealed upon photoconverting 3,3’-diaminobenzidine (DAB) into a stable, electron microscopically visible dark product. d ) Protein intensities measured with LC‒MS/MS at the input and upon enrichment of the biotinylated proteins. Two controls were used: neurons without nanobodies or neurons where an unrelated nanobody (anti-ALFA-Nb) was provided in the medium. Note that since the primary neurons do not express the ALFA tag, this control will reveal the effect of the unspecific biotinylation of the membranes occurring during the labeling period. Note that Syt1, as expected, is efficiently biotinylated and enriched upon IP with streptavidin beads. See methods for details concerning the experiments and analyses. e ) Summary of the gene ontologies (GOs; cellular components) for the proteins biotinylated upon live uptake of NbLumSyt1-APEX2 (for a detailed list, see Supplementary Table 1). As expected, synaptic components and membrane GO terms were overrepresented. f ) Possible interactors identified via live-cell proteomic mapping and enrichment vs. input and vs. IP control. Cntfr was found to be the most enriched candidate, together with other proteins that could be studied in future works. g ) Super-resolution stimulation emission depletion (STED) imaging reveals that ~ 20% of boutons labeled with live uptake are also positive for Cntfr. In this case, for cross-validation purposes, live uptake was performed with the 604.2 Syt1-luminal antibody. h ) Proximity ligation assay (in situ PLA) using antibodies against the luminal portion of Syt1 and anti-Cntfr confirmed the close proximity of these two proteins. A primary antibody against a protein not expressed in hippocampal neurons (Ribeye) was used as the negative control. i ) Blocking the network activity of primary hippocampal neurons with tetrodotoxin (TTX) for 2 h decreases the in situ PLA signal between Syt1 and Cntfr. Stimulation with the ligand of Cntfr (Cntf; 8 nM) for 2 h does not change the PLA signal between Syt1 and Cntfr. j , k ) Stimulation of neurons with Cntf for 24 h increases SV exo-endocytosis. Scale bars: 10 μm in a ; 500 nm in c ; 5 μm in g-j . The error bars indicate the means ± SEMs for panel d, and the 5th or 95th percentile for box plots; ** p < 0.01; *** p < 0.001

Figure Legend Snippet: Syt1 is found in close proximity to the ciliary neurotrophic factor receptor (Cntfr) and regulates synaptic vesicle dynamics. a-c ) NbLumSyt1-APEX2 allows efficient biotinylation of proteins in the proximity of Syt1 upon live uptake in hippocampal neurons to facilitate live-cell proteomic mapping. Representative images of neurons upon uptake of NbLumSyt1-APEX2, where biotinylated proteins are revealed with fluorescent streptavidin (a) . In the absence of H 2 O 2 , only a few endogenous biotinylated proteins are observable. In the presence of all the components, the reaction occurred efficiently, as revealed in western blot analysis of labeled neurons ( b ). To identify the interactors of Syt1, in situ proximity labeling was performed with NbLumSyt1-APEX2 ( c ). The electron microscopy image in the scheme is an example of the labeled vesicles, as revealed upon photoconverting 3,3’-diaminobenzidine (DAB) into a stable, electron microscopically visible dark product. d ) Protein intensities measured with LC‒MS/MS at the input and upon enrichment of the biotinylated proteins. Two controls were used: neurons without nanobodies or neurons where an unrelated nanobody (anti-ALFA-Nb) was provided in the medium. Note that since the primary neurons do not express the ALFA tag, this control will reveal the effect of the unspecific biotinylation of the membranes occurring during the labeling period. Note that Syt1, as expected, is efficiently biotinylated and enriched upon IP with streptavidin beads. See methods for details concerning the experiments and analyses. e ) Summary of the gene ontologies (GOs; cellular components) for the proteins biotinylated upon live uptake of NbLumSyt1-APEX2 (for a detailed list, see Supplementary Table 1). As expected, synaptic components and membrane GO terms were overrepresented. f ) Possible interactors identified via live-cell proteomic mapping and enrichment vs. input and vs. IP control. Cntfr was found to be the most enriched candidate, together with other proteins that could be studied in future works. g ) Super-resolution stimulation emission depletion (STED) imaging reveals that ~ 20% of boutons labeled with live uptake are also positive for Cntfr. In this case, for cross-validation purposes, live uptake was performed with the 604.2 Syt1-luminal antibody. h ) Proximity ligation assay (in situ PLA) using antibodies against the luminal portion of Syt1 and anti-Cntfr confirmed the close proximity of these two proteins. A primary antibody against a protein not expressed in hippocampal neurons (Ribeye) was used as the negative control. i ) Blocking the network activity of primary hippocampal neurons with tetrodotoxin (TTX) for 2 h decreases the in situ PLA signal between Syt1 and Cntfr. Stimulation with the ligand of Cntfr (Cntf; 8 nM) for 2 h does not change the PLA signal between Syt1 and Cntfr. j , k ) Stimulation of neurons with Cntf for 24 h increases SV exo-endocytosis. Scale bars: 10 μm in a ; 500 nm in c ; 5 μm in g-j . The error bars indicate the means ± SEMs for panel d, and the 5th or 95th percentile for box plots; ** p < 0.01; *** p < 0.001

Techniques Used: Western Blot, Labeling, In Situ, Electron Microscopy, Control, Membrane, Imaging, Biomarker Discovery, Proximity Ligation Assay, Negative Control, Blocking Assay, Activity Assay

other:

Article Title: Apoptosis of central and peripheral neurons can be prevented with cyclin-dependent kinase/mitogen-activated protein kinase inhibitors.
Article Snippet: Immediately after plating, survival factors were added at the following concentrations: NGF, 20 ng/ml; and ciliary neurotrophic factor (CNTF), 10 ng/ml (Alomone Laboratory, Jerusalem, Israel).

Article Title: SPARC triggers a cell-autonomous program of synapse elimination
Article Snippet: The culture medium was DMEM/F12 [1:1] containing 2.5% FBS, 2.5% rat serum (prepared in the Animal Care Facility of the Bellvitge campus of the University of Barcelona), 3–5 nM nerve growth factor (NGF), 2 nM ciliary neurotrophic factor (CNTF) (Alomone Laboratories), and 25 U/mL penicillin/streptomycin at 37 °C and 8% CO 2 .

Article Title: Survival-promoting activity of inhibitors of cyclin-dependent kinases on primary neurons correlates with inhibition of c-Jun kinase-1.
Article Snippet: Cyclin-dependent kinases and mitogen-activated protein kinases have been implicated in the regulation of cellular survival and apoptosis.. We tested the effect of two mitogen-activated/cyclindependent kinase inhibitors, olomoucine and butyrolactone I, on the in vitro survival of chick embryonic neurons.. Sensory, sympathetic, and ciliary neurons, when prepared at their respective time point of programmed cell death, could be rescued from apoptosis by both inhibitors in a dosedependent fashion.

Article Title: Inhibitors of p38 mitogen-activated protein kinase promote neuronal survival in vitro
Article Snippet: Mammalian mitogen-activated protein kinases include the extracellular signal-regulated protein kinase, the c-Jun amino-terminal kinase, and the p38 subgroups.. Sustained activation of Jun kinase and p38 have been shown to precede apoptosis of PC12 pheochromocytoma cells induced by withdrawal of trophic factors.. To investigate the possible role of p38 in neuronal apoptosis, we tested the effect of two selective p38 inhibitors, the pyridinyl imidazole compounds SB203580 and SB202190, on different populations of chick embryonic neurons in vitro.

Isolation:

Article Title: Development of Survival Responsiveness to Brain-Derived Neurotrophic Factor, Neurotrophin 3 and Neurotrophin 4/5, But Not to Nerve Growth Factor, in Cultured Motoneurons from Chick Embryo Spinal Cord
Article Snippet: Tyrosine autophosphorylation of Trks, during binding of the specific human recombinant NTs (Alomone Laboratories, Jerusalem, Israel) or after MEX stimulation was monitored by immunoblotting with the anti-phosphotyrosine monoclonal antibody 4G10 (kindly provided by Dr. David Kaplan, Montreal Neurological Institute, Montreal, Canada, or obtained from UBI, Lake Placid, NY). .. Briefly, MTNs isolated by the Nycodenz-gradient technique were plated on a substrate of poly-DL-ornithine/ laminin in 60 mm tissue culture dishes and main- tained in the presence of MEX and, in some experiments, in the presence of glial-derived neurotrophic factor (GDNF) (kindly provided by Eugene Johnson, St. Louis MO) or ciliary neurotrophic factor (CNTF) (Alomone Laboratories, Jerusalem, Israel) plus basic fibroblast growth factor (bFGF) (Boehringer Mannheim). ..

Article Title: Development of Survival Responsiveness to Brain-Derived Neurotrophic Factor, Neurotrophin 3 and Neurotrophin 4/5, But Not to Nerve Growth Factor, in Cultured Motoneurons from Chick Embryo Spinal Cord
Article Snippet: Tyrosine autophosphorylation of Trks, during binding of the specific human recombinant NTs (Alomone Laboratories, Jerusalem, Israel) or after MEX stimulation was monitored by immunoblotting with the anti-phosphotyrosine monoclonal antibody 4G10 (kindly provided by Dr. David Kaplan, Montreal Neurological Institute, Montreal, Canada, or obtained from UBI, Lake Placid, NY). .. Briefly, MTNs isolated by the Nycodenz-gradient technique were plated on a substrate of poly- dl -ornithine/laminin in 60 mm tissue culture dishes and maintained in the presence of MEX and, in some experiments, in the presence of glial-derived neurotrophic factor (GDNF) (kindly provided by Eugene Johnson, St. Louis MO) or ciliary neurotrophic factor (CNTF) (Alomone Laboratories, Jerusalem, Israel) plus basic fibroblast growth factor (bFGF) (Boehringer Mannheim). ..

Positive Control:

Article Title: Synergistic effects of osteonectin and brain-derived neurotrophic factor on axotomized retinal ganglion cells neurite outgrowth via the mitogen-activated protein kinase-extracellular signal-regulated kinase 1/2 pathways.
Article Snippet: bstract—Our previous study identified osteonectin (ON) in a creen of factors made by Schwann cells (SCs) which prooted peripheral and central neurons survival and neuritoenesis, however, the mechanisms of ON promoting effects re largely unknown.. In the present study, we investigated he effects of ON-deficient SC-conditioned medium (SCCM) nd molecular mechanisms of ON, in regulating retinal ganlion cells (RGCs) survival and neurite outgrowth.. Neonatal at RGCs and SCs were purified by immunopanning techique.

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