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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e <t>ELISA</t> quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5
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Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e ELISA quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5

Journal: Stem Cell Research & Therapy

Article Title: Targeting p75NTR activity alleviates the neurotoxic effect of high glucose on iPSC-derived dopaminergic neurons

doi: 10.1186/s13287-026-04965-y

Figure Lengend Snippet: Pro-NGF/p75NTR axis is up-regulated in glucose neurotoxicity in DA neurons. a Western blot analysis and quantification of TrkA and TrkB in Control and HG- (100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. b Western blot analysis and quantification of p75NTR in Control and HG (50mM, 100mM) treated neurons for 48 h. Protein levels were normalized to β-Actin. c Celltox cytotoxicity assay. Representative photos and quantification of dead (green) neurons after treatment with HG (50mM, 100mM) and/or p75NTR inhibitor (2.5 ng/ml, anti-p75 Receptor antibody (MC-192) Abcam) for 48 h. Scale bar 50 μm. d , e ELISA quantification of d pro-NGF protein levels in the cell lysate and e the secreted pro-NGF protein levels in the supernatant of DA neurons treated with HG (100mM) for 48 h. f Schematics of the up-regulated pro-NGF/p75NTR axis in DA neurons upon treatment with HG. For a – e data are presented as mean ± SEM of three biological replicates derived from three independent iPSC lines. For a and c statistical significance was assessed with a two-way ANOVA and Sidak’s multiple comparisons test (** P < 0.01). For b statistical significance was assessed with an ordinary one-way ANOVA and Turkey’s multiple comparisons test (** P < 0.01). For d and e statistical significance was evaluated with an unpaired t-test (* P < 0.05). Full-length blots are presented in Figure S5

Article Snippet: Pro-NGF and pro-BDNF protein levels were quantified in the cell lysis and the supernatant of neurons cultured in 12 well plate format (one well per condition) by using the human pro-NGF and the human pro-BDNF Rapid ELISA kits (Biosensis, BEK-2226-1P/2P for pro-NGF and BEK-2237-2P for pro-BDNF), respectively.

Techniques: Western Blot, Control, CellTox Assay, Cytotoxicity Assay, Enzyme-linked Immunosorbent Assay, Derivative Assay