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immunoblotting rabbit anti human trka polyclonal antibody  (Cell Signaling Technology Inc)


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    Structured Review

    Cell Signaling Technology Inc immunoblotting rabbit anti human trka polyclonal antibody
    Western blot analysis of RTK activation in A549-ACE2 cells after SARS-CoV-2 infection. Cell lysates from mock infection (Mock), A/PR8 (PR8), and SARS-CoV-2 infection at different time points were analyzed by Western blotting to detect active and total forms of EGFR ( A ), <t>TrkA</t> ( B ), and HER2 ( C ). Protein bands were quantified using Sciugo software 2.0.1. The ratio of the phosphorylated form to total form was calculated and compared to the mock (set as 1.00).
    Immunoblotting Rabbit Anti Human Trka Polyclonal Antibody, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 95/100, based on 251 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/rabbit+polyclonal+anti+trka/TrkA+Antibody/pmc12030552-51-6-12
    Average 95 stars, based on 251 article reviews
    immunoblotting rabbit anti human trka polyclonal antibody - by Bioz Stars, 2026-09
    95/100 stars

    Images

    1) Product Images from "Cellular Receptor Tyrosine Kinase Signaling Plays Important Roles in SARS-CoV-2 Infection"

    Article Title: Cellular Receptor Tyrosine Kinase Signaling Plays Important Roles in SARS-CoV-2 Infection

    Journal: Pathogens

    doi: 10.3390/pathogens14040333

    Western blot analysis of RTK activation in A549-ACE2 cells after SARS-CoV-2 infection. Cell lysates from mock infection (Mock), A/PR8 (PR8), and SARS-CoV-2 infection at different time points were analyzed by Western blotting to detect active and total forms of EGFR ( A ), TrkA ( B ), and HER2 ( C ). Protein bands were quantified using Sciugo software 2.0.1. The ratio of the phosphorylated form to total form was calculated and compared to the mock (set as 1.00).
    Figure Legend Snippet: Western blot analysis of RTK activation in A549-ACE2 cells after SARS-CoV-2 infection. Cell lysates from mock infection (Mock), A/PR8 (PR8), and SARS-CoV-2 infection at different time points were analyzed by Western blotting to detect active and total forms of EGFR ( A ), TrkA ( B ), and HER2 ( C ). Protein bands were quantified using Sciugo software 2.0.1. The ratio of the phosphorylated form to total form was calculated and compared to the mock (set as 1.00).

    Techniques Used: Western Blot, Activation Assay, Infection, Software

    Related Articles

    other:

    Article Title: NGF Induces Proliferation and Aggressiveness In Prostate Cancer Cells.
    Article Snippet: The rabbit polyclonal anti-TrkA (06-574; Millipore), or anti-ERK (C-14; Santa Cruz Biotechnology), or anti-Akt (9272; Cell Signaling) antibodies were used to detect TrkA, ERK and Akt, respectively.

    Article Title: Kai-Xin-San series formulae alleviate depressive-like behaviors on chronic mild stressed mice via regulating neurotrophic factor system on hippocampus
    Article Snippet: The primary antibodies used were: rabbit polyclonal anti-NGF (H-20, 1:1000, Santa Cruz Biotechnology), rabbit polyclonal anti-BDNF (N-20, 1:1000, Santa Cruz Biotechnology), rabbit polyclonal anti-TrkA (2505, 1:1000, Cell Signaling Technology), rabbit polyclonal anti-TrkB (4603, 1:1000, Cell Signaling Technology), mouse monoclonal anti-α tubulin (T-6074, 1:10000, Sigma-Aldrich) at 4 °C temperature overnight.

    Article Title: Nerve Growth Factor Induces Proliferation and Aggressiveness In Prostate Cancer Cells.
    Article Snippet: The rabbit polyclonal anti-TrkA (06-574; Millipore), or anti-ERK (C-14; Santa Cruz Biotechnology), or anti-Akt (9272; Cell Signaling) antibodies were used to detect TrkA, ERK and Akt, respectively.

    Article Title: Nerve Growth Factor Induces Proliferation and Aggressiveness in Prostate Cancer Cells
    Article Snippet: The rabbit polyclonal anti-TrkA (06-574; Millipore), or anti-ERK (C-14; Santa Cruz Biotechnology), or anti-Akt (9272; Cell Signaling) antibodies were used to detect TrkA, ERK and Akt, respectively.

    Western Blot:

    Article Title: GM1 Oligosaccharide Ameliorates Rett Syndrome Phenotypes In Vitro and In Vivo via Trk Receptor Activation
    Article Snippet: Secondary anti-chicken, anti-mouse, and anti-rabbit antibodies (Alexa Fluor 488, 547, and 633, respectively) were purchased from Thermo Fisher Scientific (Waltham, MA, USA). .. For WB analyses, the following antibodies were used: rabbit polyclonal anti-TOM20 (RRID: AB_2207530) purchased from ProteinTech (Manchester, UK); rabbit monoclonal anti-HTRA2 (RRID: AB_11220423), rabbit polyclonal anti-TrkA (RRID: AB_10695253), mouse monoclonal anti-β-actin (RRID:AB_2242334), rabbit monoclonal anti-TrkB (RRID:AB_2155125), rabbit monoclonal anti-phospho-TrkA Tyr674/675 /TrkB Tyr706/707 (RRID:AB_916186), and secondary horseradish peroxidase (HRP)-conjugated anti-rabbit IgG (RRID: AB_2099233) from Cell Signaling Technology (Danvers, MA, USA); mouse Oxphos cocktail (RRID: AB_2629281) from Abcam (Cambridge, UK); rabbit polyclonal anti-phospho-TrkB Tyr816 and rabbit polyclonal anti-GADPH (RRID: AB_796208) were from Merck Millipore (Burlington, MA, USA); mouse monoclonal anti-calnexin antibody (RRID: AB 397883) from BD Biosciences (San Jose, CA, USA); secondary HRP-conjugated goat anti-mouse IgG (H + L) antibody (RRID: AB_228309) from Life Technologies (Carlsbad, CA, USA); anti-HSP90 (RRID:AB_675659, and mouse monoclonal anti-phospho-TrkA Tyr490 (RRID: 628399) antibody from Santa Cruz (Dallas, TX, USA). ..

    Marker:

    Article Title: TrkA regulates the regenerative capacity of bone marrow stromal stem cells in nerve grafts
    Article Snippet: .. The following antibodies were used: mouse monoclonal anti-glial fibrillary acidic protein (GFAP, Schwann cell marker; 1:1000; Millipore, Darmstadt, Germany), mouse monoclonal anti-neurofilament 200 (NF200, axonal marker; 1:500; Boster, Wuhan, China), rabbit polyclonal anti-myelin basic protein (MBP, myelin marker; 1:500; Boster), rabbit polyclonal anti-TrkA (1:500; Millipore), rabbit polyclonal anti-TrkA phosphorylated at Tyr490 (pTrkA; 1:1000; Cell Signaling Technology, Danvers, MA, USA), rabbit polyclonal anti-Akt (1:1000; Cell Signaling Technology), rabbit polyclonal anti-Akt phosphorylated at Ser473 (pAkt; 1:1000; Cell Signaling Technology), rabbit polyclonal anti-Erk1/2 (1:1000; Cell Signaling Technology), rabbit polyclonal anti-Erk1/2 phosphorylated at Thr202/Tyr204 (pErk1/2; 1:1000; Cell Signaling Technology), rabbit polyclonal anti-Bcl-2 (anti-apoptotic protein; 1:1000; Cell Signaling Technology), rabbit polyclonal anti-Bcl-xL (anti-apoptotic protein; 1:1000; Cell Signaling Technology), rabbit polyclonal anti-Bad (pro-apoptotic factor; 1:1000; Cell Signaling Technology), rabbit polyclonal anti-Bax (pro-apoptotic factor; 1:1000; Cell Signaling Technology), rabbit polyclonal anti-cleaved caspase-3 (apoptotic marker; 1:1000; Cell Signaling Technology) and mouse monoclonal anti-β-actin (1:5000; Sigma-Aldrich). .. Secondary antibodies included goat polyclonal anti-mouse antibody conjugated with horseradish peroxidase (1:10,000; Sigma-Aldrich) and goat polyclonal anti-rabbit conjugated with horseradish peroxidase (1:10,000; Sigma-Aldrich).

    Incubation:

    Article Title: Age-induced nitrative stress decreases retrograde transport of proNGF via TrkA and increases proNGF retrograde transport and neurodegeneration via p75 NTR
    Article Snippet: .. Rat monoclonal anti-p75 NGF Receptor (Abcam; dilution 1:500) was incubated 15 h at 4°C, mouse monoclonal anti-β-Actin (Sigma-Aldrich; dilution 1:10000) was incubated for 45 min at room temperature, and rabbit polyclonal anti-TrkA (Cell Signaling Technologies; 1:500) was incubated for 2 h at room temperature and then 15 h at 4°C. .. Following three five-minute washes in TBS-T, horseradish peroxidase conjugated anti-rabbit (ThermoFisher; 1:5000), anti-mouse (Cell Signaling Technologies; 1:2000), or anti-rat (Abcam; 1:1000) secondary antibodies in 5% skim milk in TBS-T were incubated for 1 h at room temperature.

    Article Title: Age-induced nitrative stress decreases retrograde transport of proNGF via TrkA and increases proNGF retrograde transport and neurodegeneration via p75 NTR .
    Article Snippet: .. Rat monoclonal anti-p75 NGF Receptor (Abcam; dilution 1:500) was incubated 15 h at 4°C, mouse monoclonal anti-βActin (Sigma-Aldrich; dilution 1:10000) was incubated for 45 min at room temperature, and rabbit polyclonal anti-TrkA (Cell Signaling Technologies; 1:500) was incubated for 2 h at room temperature and then 15 h at 4°C. .. Following three five-minute washes in TBS-T, horseradish peroxidase conjugated anti-rabbit (ThermoFisher; 1:5000), anti-mouse (Cell Signaling Technologies; 1:2000), or anti-rat (Abcam; 1:1000) secondary antibodies in 5% skim milk in TBS-T were incubated for 1 h at room temperature.



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    (A-C’) Representative transverse sections through the trigeminal ganglion of an E5.5/HH28 chick embryo following immunohistochemistry <t>for</t> <t>N-cadherin</t> (N-cad) and <t>TrkA.</t> Scale bar in (A) is 50 µm and applies to (B, C). Scale bar in (A’) is 50 µm and applies to (B’, C’). (D-I) Representative trigeminal ganglia explant cultures after immunocytochemistry for N-cadherin and TrkA. Scale bar in (D) is 25 µm and applies to (E-I). Arrowheads indicate colocalization of N-cadherin and TrkA. (J, K) Immunoblots of HH28-30 trigeminal ganglia for TrkA with lysate from the embryo ( in vivo ) (J) and from trigeminal ganglia explant cultures ( ex vivo ) (K). Pie charts represent percentage of total signal coming from the different bands of TrkA.
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    Cell Signaling Technology Inc polyclonal rabbit anti human y490 phosphorylated trka
    ( a ) Indirect IFs demonstrating increased overlapping (yellow/orange) immunoreactivity for <t>TrkA</t> (green) and MitoTracker-labeled mitochondria (red) (upper panels) and <t>Y490</t> <t>phosphorylated</t> TrkAIII (pTrkAIII, green) and MitoTracker-labeled mitochondria (red) (lower panels) in DTT-treated (5 mM for 6 h) TrkAIII SH-SY5Y cells compared to untreated controls (control). DAPI stained nuclei are blue. (bar = 50 μm). ( b ) Western blots demonstrating TrkAIII cleavage and Y674/5 phosphorylation in mitochondria (50 mg) from DTT-treated (5 mM for 6 h) TrkAIII SH-SY5Y cells compared to untreated TrkAIII SH-SY5Y controls. ( c ) Western blots demonstrating increased PTPase oxidation (arrows) in mitochondria (50 mg) from DTT-treated TrkAIII SH-SY5Y cells (5 mM for 6 h) compared to untreated controls (Con). ( d ) Phase contrast images merged with green fluorescence and histogram demonstrating significant differences (* p < 0.0001) in pcDNA-SH-SY5Y and TrkAIII SH-SY5Y percentage cell death at 24 h and 48 h, induced by 5 mM DTT.
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    Image Search Results


    HG-induced TrkA phosphorylation is associated with podocyte inflammation and injury. (A–D) Representative western blots show the levels of (A) TrkA and phospho-TrkA (Tyr490); (B) ERK, phospho-ERK and EGR1; (C) p35 and CDK5; and (D) IL-1β and TNF-α in MPC5 cells from the LG, HG, and LG + Man groups at 24 h (E) Representative immunofluorescence double staining shows the expression of phospho-TrkA (Tyr490) and synaptopodin in MPC5 cells from the LG, HG, and LG + Man groups. ****p < 0.0001. p < 0.05 was considered statistically significant; ns indicates not significant.

    Journal: Frontiers in Endocrinology

    Article Title: Unveiling the TrkA-p35/CDK5 axis: a novel therapeutic target in diabetic kidney disease

    doi: 10.3389/fendo.2026.1791283

    Figure Lengend Snippet: HG-induced TrkA phosphorylation is associated with podocyte inflammation and injury. (A–D) Representative western blots show the levels of (A) TrkA and phospho-TrkA (Tyr490); (B) ERK, phospho-ERK and EGR1; (C) p35 and CDK5; and (D) IL-1β and TNF-α in MPC5 cells from the LG, HG, and LG + Man groups at 24 h (E) Representative immunofluorescence double staining shows the expression of phospho-TrkA (Tyr490) and synaptopodin in MPC5 cells from the LG, HG, and LG + Man groups. ****p < 0.0001. p < 0.05 was considered statistically significant; ns indicates not significant.

    Article Snippet: Anti-phospho-TrkA (pTyr490) antibody (Cat. No. RA18018-100) was purchased from Origene (Rockville, MD, USA).

    Techniques: Phospho-proteomics, Western Blot, Immunofluorescence, Double Staining, Expressing

    TrkA inhibition reduces the levels of p-TrkA, p-ERK, EGR1, p35, and CDK5 in the kidneys of db/db mice. (A–C) Representative western blots show the protein expression levels of (A) TrkA and phospho-TrkA (Tyr490), (B) ERK, p-ERK and EGR1, and (C) p35 and CDK5 in the kidney samples from the control, db/db, Inh-NC, and Inh-TrkA groups of mice. (D) Representative IF images show the expression of Synaptopodin (red) in the glomeruli of control, db/db, Inh-NC, and Inh-TrkA groups of mice (400X). ***p < 0.001. p < 0.05 was considered statistically significant; ns indicates not significant.

    Journal: Frontiers in Endocrinology

    Article Title: Unveiling the TrkA-p35/CDK5 axis: a novel therapeutic target in diabetic kidney disease

    doi: 10.3389/fendo.2026.1791283

    Figure Lengend Snippet: TrkA inhibition reduces the levels of p-TrkA, p-ERK, EGR1, p35, and CDK5 in the kidneys of db/db mice. (A–C) Representative western blots show the protein expression levels of (A) TrkA and phospho-TrkA (Tyr490), (B) ERK, p-ERK and EGR1, and (C) p35 and CDK5 in the kidney samples from the control, db/db, Inh-NC, and Inh-TrkA groups of mice. (D) Representative IF images show the expression of Synaptopodin (red) in the glomeruli of control, db/db, Inh-NC, and Inh-TrkA groups of mice (400X). ***p < 0.001. p < 0.05 was considered statistically significant; ns indicates not significant.

    Article Snippet: Anti-phospho-TrkA (pTyr490) antibody (Cat. No. RA18018-100) was purchased from Origene (Rockville, MD, USA).

    Techniques: Inhibition, Western Blot, Expressing, Control

    (A-C’) Representative transverse sections through the trigeminal ganglion of an E5.5/HH28 chick embryo following immunohistochemistry for N-cadherin (N-cad) and TrkA. Scale bar in (A) is 50 µm and applies to (B, C). Scale bar in (A’) is 50 µm and applies to (B’, C’). (D-I) Representative trigeminal ganglia explant cultures after immunocytochemistry for N-cadherin and TrkA. Scale bar in (D) is 25 µm and applies to (E-I). Arrowheads indicate colocalization of N-cadherin and TrkA. (J, K) Immunoblots of HH28-30 trigeminal ganglia for TrkA with lysate from the embryo ( in vivo ) (J) and from trigeminal ganglia explant cultures ( ex vivo ) (K). Pie charts represent percentage of total signal coming from the different bands of TrkA.

    Journal: bioRxiv

    Article Title: Multiple glycoforms of TrkA interact with N-cadherin during trigeminal ganglion neurodevelopment

    doi: 10.1101/2025.11.14.688512

    Figure Lengend Snippet: (A-C’) Representative transverse sections through the trigeminal ganglion of an E5.5/HH28 chick embryo following immunohistochemistry for N-cadherin (N-cad) and TrkA. Scale bar in (A) is 50 µm and applies to (B, C). Scale bar in (A’) is 50 µm and applies to (B’, C’). (D-I) Representative trigeminal ganglia explant cultures after immunocytochemistry for N-cadherin and TrkA. Scale bar in (D) is 25 µm and applies to (E-I). Arrowheads indicate colocalization of N-cadherin and TrkA. (J, K) Immunoblots of HH28-30 trigeminal ganglia for TrkA with lysate from the embryo ( in vivo ) (J) and from trigeminal ganglia explant cultures ( ex vivo ) (K). Pie charts represent percentage of total signal coming from the different bands of TrkA.

    Article Snippet: The membrane was blocked in 5% milk (in 1X PBS+0.1% Tween-20 (PTW)) for 30 minutes at room temperature followed by overnight incubation at 4°C with the following primary antibodies diluted in blocking solution: N-cadherin (MNCD2, DSHB 1:100) and TrkA (MSU, 1:5000 or Origene, 1:1000).

    Techniques: Immunohistochemistry, Immunocytochemistry, Western Blot, In Vivo, Ex Vivo

    (A) Dissected trigeminal ganglion from an E5.5/HH28 chick embryo. Dotted lines indicate approximate location where cuts were made to separate trigeminal cell bodies from axons. Scale bar is 1mm. (B-C) N-cadherin-TrkA co-immunoprecipitation on pooled HH28-30 trigeminal cell bodies and axons. Input lanes contain 10% of lysate loaded in immunoprecipitation lanes. OpV, ophthalmic; MmV, maxillomandibular; IP, immunoprecipitation; IgG, Immunoglobulin G. N=3.

    Journal: bioRxiv

    Article Title: Multiple glycoforms of TrkA interact with N-cadherin during trigeminal ganglion neurodevelopment

    doi: 10.1101/2025.11.14.688512

    Figure Lengend Snippet: (A) Dissected trigeminal ganglion from an E5.5/HH28 chick embryo. Dotted lines indicate approximate location where cuts were made to separate trigeminal cell bodies from axons. Scale bar is 1mm. (B-C) N-cadherin-TrkA co-immunoprecipitation on pooled HH28-30 trigeminal cell bodies and axons. Input lanes contain 10% of lysate loaded in immunoprecipitation lanes. OpV, ophthalmic; MmV, maxillomandibular; IP, immunoprecipitation; IgG, Immunoglobulin G. N=3.

    Article Snippet: The membrane was blocked in 5% milk (in 1X PBS+0.1% Tween-20 (PTW)) for 30 minutes at room temperature followed by overnight incubation at 4°C with the following primary antibodies diluted in blocking solution: N-cadherin (MNCD2, DSHB 1:100) and TrkA (MSU, 1:5000 or Origene, 1:1000).

    Techniques: Immunoprecipitation

    (A) Immunoblots of E6.5/HH28-30 trigeminal ganglia lysate following Mem-PER membrane extraction protocol for TrkA, N-cadherin, Tubb3, GM130 (Golgi protein), and SERCA2 (ER protein) using membrane-extracted and cytosolic-extracted protein fractions. (B) N-cadherin-TrkA co-immunoprecipitation on membrane-extracted fraction. Input contains 10% of lysate loaded in immunoprecipitation lanes. The top blot image is from one experiment using a high exposure and the bottom blot image is from a separate experiment with a lower exposure. IP, immunoprecipitation, IgG, Immunoglobulin G (N=3).

    Journal: bioRxiv

    Article Title: Multiple glycoforms of TrkA interact with N-cadherin during trigeminal ganglion neurodevelopment

    doi: 10.1101/2025.11.14.688512

    Figure Lengend Snippet: (A) Immunoblots of E6.5/HH28-30 trigeminal ganglia lysate following Mem-PER membrane extraction protocol for TrkA, N-cadherin, Tubb3, GM130 (Golgi protein), and SERCA2 (ER protein) using membrane-extracted and cytosolic-extracted protein fractions. (B) N-cadherin-TrkA co-immunoprecipitation on membrane-extracted fraction. Input contains 10% of lysate loaded in immunoprecipitation lanes. The top blot image is from one experiment using a high exposure and the bottom blot image is from a separate experiment with a lower exposure. IP, immunoprecipitation, IgG, Immunoglobulin G (N=3).

    Article Snippet: The membrane was blocked in 5% milk (in 1X PBS+0.1% Tween-20 (PTW)) for 30 minutes at room temperature followed by overnight incubation at 4°C with the following primary antibodies diluted in blocking solution: N-cadherin (MNCD2, DSHB 1:100) and TrkA (MSU, 1:5000 or Origene, 1:1000).

    Techniques: Western Blot, Membrane, Extraction, Immunoprecipitation

    Western blot analysis of RTK activation in A549-ACE2 cells after SARS-CoV-2 infection. Cell lysates from mock infection (Mock), A/PR8 (PR8), and SARS-CoV-2 infection at different time points were analyzed by Western blotting to detect active and total forms of EGFR ( A ), TrkA ( B ), and HER2 ( C ). Protein bands were quantified using Sciugo software 2.0.1. The ratio of the phosphorylated form to total form was calculated and compared to the mock (set as 1.00).

    Journal: Pathogens

    Article Title: Cellular Receptor Tyrosine Kinase Signaling Plays Important Roles in SARS-CoV-2 Infection

    doi: 10.3390/pathogens14040333

    Figure Lengend Snippet: Western blot analysis of RTK activation in A549-ACE2 cells after SARS-CoV-2 infection. Cell lysates from mock infection (Mock), A/PR8 (PR8), and SARS-CoV-2 infection at different time points were analyzed by Western blotting to detect active and total forms of EGFR ( A ), TrkA ( B ), and HER2 ( C ). Protein bands were quantified using Sciugo software 2.0.1. The ratio of the phosphorylated form to total form was calculated and compared to the mock (set as 1.00).

    Article Snippet: The following antibodies were used for immunoblotting: rabbit anti-human TrkA polyclonal antibody (Cell Signaling Technology, 2505S, Danvers, MA, USA), rabbit anti-human pTrkA monoclonal antibody (mAb) (Cell Signaling Tech, 4168S), rabbit anti-human HER2 mAb (Cell Signaling Tech, 2165S), rabbit anti-human pHER2 mAb (Cell Signaling Tech, 2243L), mouse anti-human EGFR mAb (Cell Signaling Tech, 2239S), rabbit anti-human pEGFR mAb (Cell Signaling Tech, 3777S), rabbit anti-human GAPDH mAb (Cell Signaling Tech, 2118S), anti-rabbit HRP conjugated (Bio Rad, 1706515, Hercules, CA, USA), and anti-mouse IgG HRP conjugated (R&D, HAF007).

    Techniques: Western Blot, Activation Assay, Infection, Software

    ( a ) Indirect IFs demonstrating increased overlapping (yellow/orange) immunoreactivity for TrkA (green) and MitoTracker-labeled mitochondria (red) (upper panels) and Y490 phosphorylated TrkAIII (pTrkAIII, green) and MitoTracker-labeled mitochondria (red) (lower panels) in DTT-treated (5 mM for 6 h) TrkAIII SH-SY5Y cells compared to untreated controls (control). DAPI stained nuclei are blue. (bar = 50 μm). ( b ) Western blots demonstrating TrkAIII cleavage and Y674/5 phosphorylation in mitochondria (50 mg) from DTT-treated (5 mM for 6 h) TrkAIII SH-SY5Y cells compared to untreated TrkAIII SH-SY5Y controls. ( c ) Western blots demonstrating increased PTPase oxidation (arrows) in mitochondria (50 mg) from DTT-treated TrkAIII SH-SY5Y cells (5 mM for 6 h) compared to untreated controls (Con). ( d ) Phase contrast images merged with green fluorescence and histogram demonstrating significant differences (* p < 0.0001) in pcDNA-SH-SY5Y and TrkAIII SH-SY5Y percentage cell death at 24 h and 48 h, induced by 5 mM DTT.

    Journal: International Journal of Molecular Sciences

    Article Title: Molecular Characterization and Inhibition of a Novel Stress-Induced Mitochondrial Protecting Role for Misfolded TrkAIII in Human SH-SY5Y Neuroblastoma Cells

    doi: 10.3390/ijms25105475

    Figure Lengend Snippet: ( a ) Indirect IFs demonstrating increased overlapping (yellow/orange) immunoreactivity for TrkA (green) and MitoTracker-labeled mitochondria (red) (upper panels) and Y490 phosphorylated TrkAIII (pTrkAIII, green) and MitoTracker-labeled mitochondria (red) (lower panels) in DTT-treated (5 mM for 6 h) TrkAIII SH-SY5Y cells compared to untreated controls (control). DAPI stained nuclei are blue. (bar = 50 μm). ( b ) Western blots demonstrating TrkAIII cleavage and Y674/5 phosphorylation in mitochondria (50 mg) from DTT-treated (5 mM for 6 h) TrkAIII SH-SY5Y cells compared to untreated TrkAIII SH-SY5Y controls. ( c ) Western blots demonstrating increased PTPase oxidation (arrows) in mitochondria (50 mg) from DTT-treated TrkAIII SH-SY5Y cells (5 mM for 6 h) compared to untreated controls (Con). ( d ) Phase contrast images merged with green fluorescence and histogram demonstrating significant differences (* p < 0.0001) in pcDNA-SH-SY5Y and TrkAIII SH-SY5Y percentage cell death at 24 h and 48 h, induced by 5 mM DTT.

    Article Snippet: Polyclonal rabbit anti-human Y490 phosphorylated TrkA (9141), polyclonal rabbit anti-human Akt (9272), polyclonal rabbit anti-human phospho-Ser 473-Akt (4060), polyclonal rabbit anti-human Y674/675 (4621), antibody was from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Labeling, Control, Staining, Western Blot, Phospho-proteomics, Fluorescence

    ( a ) Western blots demonstrating TrkAIII cleavage and phosphorylation in mitochondria (50 μg) from TrkAIII SH-SY5Y cells treated with 5 mM DTT alone for 6 h and in mitochondria from TrkAIII SH-SY5Y cells co-treated with 5 mM DTT and either HA-15 (20 μM) (DTT/HA); brefeldin A (5 mg/mL) (DTT/BfA) or W7 (60 μM) (DTT/W7) plus non-phosphorylated TrkAIII in mitochondria from untreated TrkAIII SH-SY5Y cells (Con) and lack of TrkA or phosphorylated TrkA immunoreactivity in mitochondria (50 μg) from untreated (Con) and DTT-treated (5 mm for 6 h) pcDNA-SH-SY5Y cells. ( b ) Line graphs demonstrating significant inhibition (*) of pcDNA-SH-SY5Y and TrkAIII SH-SY5Y proliferation by BfA (5 μg/mL) and W7 (60 μM) but not by HA-15 (20 μM) at 24 and 48 h (* p < 0.0001). ( c ) Phase contrast images merged with green fluorescence plus histograms demonstrating percentage pcDNA-SH-SY5Y and TrkAIII SH-SY5Y cell death induced by 5 mM DTT alone (DTT), DTT and either HA-15 (20 μM) (DTT/HA), BfA (5 μg/mL) (DTT/BfA) or W7 (60 μM) (DTT/W7), plus pcDNA-SH-SY5Y and TrkAIII SH-SY5Y cell death induced by W7 (60 μM) alone (W7) at 24 and 48 h (* p < 0.006).

    Journal: International Journal of Molecular Sciences

    Article Title: Molecular Characterization and Inhibition of a Novel Stress-Induced Mitochondrial Protecting Role for Misfolded TrkAIII in Human SH-SY5Y Neuroblastoma Cells

    doi: 10.3390/ijms25105475

    Figure Lengend Snippet: ( a ) Western blots demonstrating TrkAIII cleavage and phosphorylation in mitochondria (50 μg) from TrkAIII SH-SY5Y cells treated with 5 mM DTT alone for 6 h and in mitochondria from TrkAIII SH-SY5Y cells co-treated with 5 mM DTT and either HA-15 (20 μM) (DTT/HA); brefeldin A (5 mg/mL) (DTT/BfA) or W7 (60 μM) (DTT/W7) plus non-phosphorylated TrkAIII in mitochondria from untreated TrkAIII SH-SY5Y cells (Con) and lack of TrkA or phosphorylated TrkA immunoreactivity in mitochondria (50 μg) from untreated (Con) and DTT-treated (5 mm for 6 h) pcDNA-SH-SY5Y cells. ( b ) Line graphs demonstrating significant inhibition (*) of pcDNA-SH-SY5Y and TrkAIII SH-SY5Y proliferation by BfA (5 μg/mL) and W7 (60 μM) but not by HA-15 (20 μM) at 24 and 48 h (* p < 0.0001). ( c ) Phase contrast images merged with green fluorescence plus histograms demonstrating percentage pcDNA-SH-SY5Y and TrkAIII SH-SY5Y cell death induced by 5 mM DTT alone (DTT), DTT and either HA-15 (20 μM) (DTT/HA), BfA (5 μg/mL) (DTT/BfA) or W7 (60 μM) (DTT/W7), plus pcDNA-SH-SY5Y and TrkAIII SH-SY5Y cell death induced by W7 (60 μM) alone (W7) at 24 and 48 h (* p < 0.006).

    Article Snippet: Polyclonal rabbit anti-human Y490 phosphorylated TrkA (9141), polyclonal rabbit anti-human Akt (9272), polyclonal rabbit anti-human phospho-Ser 473-Akt (4060), polyclonal rabbit anti-human Y674/675 (4621), antibody was from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Western Blot, Phospho-proteomics, Inhibition, Fluorescence