hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc hpk1
    Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc hpk1
    Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    anti hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anti hpk1
    Anti Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc hpk1
    Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc hpk1
    a , The relative quantitation of proteins of interest immunoprecipitated by bead-conjugated AKT phospho-substrate antibody in (n=2 from two independent experiments). b , MEG NBCs and GCBCs were pretreated with AKT inhibitor for 10 min and then stimulated with anti-IgM for 5 min or left unstimulated. Cell lysates were immunoprecipitated by AKT phospho-substrate antibody, and then probed by immunoblot for each of the indicated proteins. Data represent one of two independent experiments. c , In vitro kinase reactions were performed for 30 min by incubating recombinant pre-activated AKT with CSK, SHP-1, <t>HPK1</t> or LYN as substrates. Immunoblot using the AKT phospho-substrate antibody was utilized to monitor phosphorylation of the target proteins. A representative of two independent experiments is shown.
    Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "The AKT kinase signaling network is rewired by PTEN to control proximal BCR signaling in germinal center B cells"

    Article Title: The AKT kinase signaling network is rewired by PTEN to control proximal BCR signaling in germinal center B cells

    Journal: Nature immunology

    doi: 10.1038/s41590-019-0376-3

    a , The relative quantitation of proteins of interest immunoprecipitated by bead-conjugated AKT phospho-substrate antibody in (n=2 from two independent experiments). b , MEG NBCs and GCBCs were pretreated with AKT inhibitor for 10 min and then stimulated with anti-IgM for 5 min or left unstimulated. Cell lysates were immunoprecipitated by AKT phospho-substrate antibody, and then probed by immunoblot for each of the indicated proteins. Data represent one of two independent experiments. c , In vitro kinase reactions were performed for 30 min by incubating recombinant pre-activated AKT with CSK, SHP-1, HPK1 or LYN as substrates. Immunoblot using the AKT phospho-substrate antibody was utilized to monitor phosphorylation of the target proteins. A representative of two independent experiments is shown.
    Figure Legend Snippet: a , The relative quantitation of proteins of interest immunoprecipitated by bead-conjugated AKT phospho-substrate antibody in (n=2 from two independent experiments). b , MEG NBCs and GCBCs were pretreated with AKT inhibitor for 10 min and then stimulated with anti-IgM for 5 min or left unstimulated. Cell lysates were immunoprecipitated by AKT phospho-substrate antibody, and then probed by immunoblot for each of the indicated proteins. Data represent one of two independent experiments. c , In vitro kinase reactions were performed for 30 min by incubating recombinant pre-activated AKT with CSK, SHP-1, HPK1 or LYN as substrates. Immunoblot using the AKT phospho-substrate antibody was utilized to monitor phosphorylation of the target proteins. A representative of two independent experiments is shown.

    Techniques Used: Quantitation Assay, Immunoprecipitation, Western Blot, In Vitro, Recombinant

    a , Mass spectrometry was performed on the in vitro kinase reaction of AKT and CSK to identify the AKT phosphorylation site on CSK (S284). S284 phosphorylation was modeled onto the crystal structure of CSK bound to C-SRC (PDB ID 3D7T). b , unphosphorylated CSK or AKT-phosphorylated CSK was reacted with LYN and immunoblot was utilized to monitor LYN Y507 phosphorylation. Densitometry was utilized to quantitate LYN Y507 phosphorylation (n=3 from three independent experiments). The resulting kinetic traces, shown are means ± SD, were fit with a linear model to determine the relative reaction rate for LYN phosphorylation. P values were derived with a two-tailed Student’s t -test (t=21.55, d.f.=4). c , Mass spectrometry was utilized to map the AKT phosphorylation site on SHP-1 to T394, which is in the catalytic domain. d , A phosphatase assay was performed to measure the activity of unphosphorylated SHP-1 and AKT-phosphorylated SHP-1. The resulting kinetic traces were fit with a linear model to determine the relative reaction rates (n=3 from three independent experiments). Shown are means ± SD, P values were calculated with a two-tailed Student’s t -test (t=4.963, d.f.=4). e , Unphosphorylated HPK1 and AKT-phosphorylated HPK1 were incubated with recombinant BLNK and immunoblot was performed with an antibody against phospho-threonine residue. Densitometry was utilized to quantitate BLNK phosphorylation (n=2 from two independent experiments). Shown are means ± SD. P values, two-way ANOVA with Sidak’s multiple comparisons test (F=15.95, d.f.=5).
    Figure Legend Snippet: a , Mass spectrometry was performed on the in vitro kinase reaction of AKT and CSK to identify the AKT phosphorylation site on CSK (S284). S284 phosphorylation was modeled onto the crystal structure of CSK bound to C-SRC (PDB ID 3D7T). b , unphosphorylated CSK or AKT-phosphorylated CSK was reacted with LYN and immunoblot was utilized to monitor LYN Y507 phosphorylation. Densitometry was utilized to quantitate LYN Y507 phosphorylation (n=3 from three independent experiments). The resulting kinetic traces, shown are means ± SD, were fit with a linear model to determine the relative reaction rate for LYN phosphorylation. P values were derived with a two-tailed Student’s t -test (t=21.55, d.f.=4). c , Mass spectrometry was utilized to map the AKT phosphorylation site on SHP-1 to T394, which is in the catalytic domain. d , A phosphatase assay was performed to measure the activity of unphosphorylated SHP-1 and AKT-phosphorylated SHP-1. The resulting kinetic traces were fit with a linear model to determine the relative reaction rates (n=3 from three independent experiments). Shown are means ± SD, P values were calculated with a two-tailed Student’s t -test (t=4.963, d.f.=4). e , Unphosphorylated HPK1 and AKT-phosphorylated HPK1 were incubated with recombinant BLNK and immunoblot was performed with an antibody against phospho-threonine residue. Densitometry was utilized to quantitate BLNK phosphorylation (n=2 from two independent experiments). Shown are means ± SD. P values, two-way ANOVA with Sidak’s multiple comparisons test (F=15.95, d.f.=5).

    Techniques Used: Mass Spectrometry, In Vitro, Western Blot, Derivative Assay, Two Tailed Test, Phosphatase Assay, Activity Assay, Incubation, Recombinant

    Purified MEG NBCs and GCBCs were incubated with PTEN inhibitor (SF1670) or DMSO for 30 min and then stimulated with anti-IgM for 5 min. The proteomic study workflow described in was utilized to identify AKT substrates (n=2 from two independent experiments). a , The PANTHER software package was utilized to identify pathways targeted by AKT in GCBCs. Heat map shows the relative protein abundance by normalizing the peak area for each experimental group to the maximum peak area observed for each protein. b , The peak area from the mass spectrometric assay was calculated for CSK, SHP1, HPK1 and ARP2. Bar graphs show the mean of the results with dots showing results of each independent experiment.
    Figure Legend Snippet: Purified MEG NBCs and GCBCs were incubated with PTEN inhibitor (SF1670) or DMSO for 30 min and then stimulated with anti-IgM for 5 min. The proteomic study workflow described in was utilized to identify AKT substrates (n=2 from two independent experiments). a , The PANTHER software package was utilized to identify pathways targeted by AKT in GCBCs. Heat map shows the relative protein abundance by normalizing the peak area for each experimental group to the maximum peak area observed for each protein. b , The peak area from the mass spectrometric assay was calculated for CSK, SHP1, HPK1 and ARP2. Bar graphs show the mean of the results with dots showing results of each independent experiment.

    Techniques Used: Purification, Incubation, Software

    hpk 1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc hpk 1
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    hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc hpk1
    Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc hpk1
    Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    anti hpk1  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anti hpk1
    Anti Hpk1, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    anti hpk1  (Cell Signaling Technology Inc)


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