Western Blot:Article Title: Dynein-Mediated Trafficking: A New Mechanism of Diabetic Podocytopathy
Article Snippet: Table 5 Plasmids Company Catalog No. pCDNA3 mouse nephrin Gift from Dr. Lawrence B. Holzman GFP-Rab7 Addgene 12605 GFP-Rab11 Addgene 12674 pEGFP-DCTN1 Addgene 36154 pCMV3-C-GFPSpark-Dynll1 SinoBiochemical MG51246-ACG EB3-tdTomato Addgene 50708 Open in a separate window Plasmids. .. Table 2 Antibody Dilution Company Cat# Mouse antinephrin (G-8) IF: 1:100 Santa Cruz sc-376522 Mouse antinephrin conjugated to agarose IP: 2 μg/400 μl lysate Santa Cruz sc-376522 AC Normal mouse IgG conjugated to agarose IP: 2 μg/400 μl lysate Santa Cruz sc-2343 Rabbit antinephrin WB: 1:1000 Invitrogen PA5-91907 Sheep anti-DCTN1 WB: 1:1000 IF: 1:100 R&D Systems AF6657 Mouse anti-Dynll1 IF: 1:100 Santa Cruz sc-136287 Rabbit anti-Dynll1 WB: 1:1000 IF: 1:100 Thermo Fisher PA5-97920 Rabbit anti-INF2 IF: 1:100 Bethyl Lab A303-427A Rabbit anti-HDAC6 WB: 1:1000 Novus NBP1-78981 Donkey anti-mouse (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32766 Donkey anti-rabbit (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32790 Donkey anti-mouse (Alexa Fluor 594) IF: 1:200 Thermo Fisher A32744 Donkey anti-sheep (Alexa Fluor 488) IF: 1:200 Thermo Fisher A-11015 Mouse anti-rabbit IgG-HRP WB: 1:5000 Santa Cruz sc-2357 Rabbit anti-sheep IgG-HRP WB: 1:5000 Thermo Fisher 31480 Biotin mouse anti-ubiquitin WB: 1:1000 Invitrogen 13-6078-82 Mouse anti β-actin-HRP WB: 1:5000 Santa Cruz sc-47778 Mouse anti-WT1 IF: 1:100 Novus NBP24460700 Mouse anti-acetylated α-tubulin IF: 1:200 Santa Cruz sc-23950 Acti-stain 488 phalloidin Cytoskeleton PHDG1-A ViaFluor microtubule stains (488) Biotium 70062 Open in a separate window IF, immunofluorescent; IP, immunoprecipitation; WB, Western blotting; INF2, inverted formin 2; HDAC6, histone deacetylase 6; HRP, horseradish peroxidase. ..
Article Title: Dynein-Mediated Trafficking: A New Mechanism of Diabetic Podocytopathy
Article Snippet: .. Antibodies and fluorescent probes (immunofluorescent, Western blotting, immunoprecipitation] Antibody Dilution Company Cat# Mouse antinephrin (G-8) IF: 1:100 Santa Cruz sc-376522 Mouse antinephrin conjugated to agarose IP: 2 mg/400 ml lysate Santa Cruz sc-376522 AC Normal mouse IgG conjugated to agarose IP: 2 mg/400 ml lysate Santa Cruz sc-2343 Rabbit antinephrin WB: 1:1000 Invitrogen PA5-91907 Sheep anti-DCTN1 WB: 1:1000 IF: 1:100 R&D Systems AF6657 Mouse anti-Dynll1 IF: 1:100 Santa Cruz sc-136287 Rabbit anti-Dynll1 WB: 1:1000 IF: 1:100 Thermo Fisher PA5-97920 Rabbit anti-INF2 IF: 1:100 Bethyl Lab A303-427A Rabbit anti-HDAC6 WB: 1:1000 Novus NBP1-78981 Donkey anti-mouse (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32766 Donkey anti-rabbit (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32790 Donkey anti-mouse (Alexa Fluor 594) IF: 1:200 Thermo Fisher A32744 Donkey anti-sheep (Alexa Fluor 488) IF: 1:200 Thermo Fisher A-11015 Mouse anti-rabbit IgG-HRP WB: 1:5000 Santa Cruz sc-2357 Rabbit anti-sheep IgG-HRP WB: 1:5000 Thermo Fisher 31480 Biotin mouse anti-ubiquitin WB: 1:1000 Invitrogen 13-6078-82 Mouse anti b-actin-HRP WB: 1:5000 Santa Cruz sc-47778 Mouse anti-WT1 IF: 1:100 Novus NBP24460700 Mouse anti-acetylated a-tubulin IF: 1:200 Santa Cruz sc-23950 Acti-stain 488 phalloidin Cytoskeleton PHDG1-A ViaFluor microtubule stains (488) Biotium 70062 IF, immunofluorescent; IP, immunoprecipitation; WB, Western blotting; INF2, inverted formin 2; HDAC6, histone deacetylase 6; HRP, horseradish peroxidase. .. 164 KIDNEY360 D ow nladed fom http:urnals.lw w .om /kidey360 by B hD M feP H K a1zE um tQ fN a+ kLhE Z gsIH 4X M ihC yw C X 1A W nY Q p/IlQ rH D 3i3D 0O dR yi7T vS F 4C 3V C 1ybggQ Z X gG 2M w lZ LeI= on 018/2024 Co-Immunoprecipitation and Immunoblotting Cultured podocytes were lysed on ice for 1 hour in Nonidet P40 lysis buffer with 0.5% deoxycholate, cOmplete, Mini Protease Inhibitor Cocktail (Roche, #11836153001), and PhosSTOP phosphatase inhibitor cocktail (Sigma, #4906845001).
Immunoprecipitation:Article Title: Dynein-Mediated Trafficking: A New Mechanism of Diabetic Podocytopathy
Article Snippet: Table 5 Plasmids Company Catalog No. pCDNA3 mouse nephrin Gift from Dr. Lawrence B. Holzman GFP-Rab7 Addgene 12605 GFP-Rab11 Addgene 12674 pEGFP-DCTN1 Addgene 36154 pCMV3-C-GFPSpark-Dynll1 SinoBiochemical MG51246-ACG EB3-tdTomato Addgene 50708 Open in a separate window Plasmids. .. Table 2 Antibody Dilution Company Cat# Mouse antinephrin (G-8) IF: 1:100 Santa Cruz sc-376522 Mouse antinephrin conjugated to agarose IP: 2 μg/400 μl lysate Santa Cruz sc-376522 AC Normal mouse IgG conjugated to agarose IP: 2 μg/400 μl lysate Santa Cruz sc-2343 Rabbit antinephrin WB: 1:1000 Invitrogen PA5-91907 Sheep anti-DCTN1 WB: 1:1000 IF: 1:100 R&D Systems AF6657 Mouse anti-Dynll1 IF: 1:100 Santa Cruz sc-136287 Rabbit anti-Dynll1 WB: 1:1000 IF: 1:100 Thermo Fisher PA5-97920 Rabbit anti-INF2 IF: 1:100 Bethyl Lab A303-427A Rabbit anti-HDAC6 WB: 1:1000 Novus NBP1-78981 Donkey anti-mouse (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32766 Donkey anti-rabbit (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32790 Donkey anti-mouse (Alexa Fluor 594) IF: 1:200 Thermo Fisher A32744 Donkey anti-sheep (Alexa Fluor 488) IF: 1:200 Thermo Fisher A-11015 Mouse anti-rabbit IgG-HRP WB: 1:5000 Santa Cruz sc-2357 Rabbit anti-sheep IgG-HRP WB: 1:5000 Thermo Fisher 31480 Biotin mouse anti-ubiquitin WB: 1:1000 Invitrogen 13-6078-82 Mouse anti β-actin-HRP WB: 1:5000 Santa Cruz sc-47778 Mouse anti-WT1 IF: 1:100 Novus NBP24460700 Mouse anti-acetylated α-tubulin IF: 1:200 Santa Cruz sc-23950 Acti-stain 488 phalloidin Cytoskeleton PHDG1-A ViaFluor microtubule stains (488) Biotium 70062 Open in a separate window IF, immunofluorescent; IP, immunoprecipitation; WB, Western blotting; INF2, inverted formin 2; HDAC6, histone deacetylase 6; HRP, horseradish peroxidase. ..
Article Title: Dynein-Mediated Trafficking: A New Mechanism of Diabetic Podocytopathy
Article Snippet: .. Antibodies and fluorescent probes (immunofluorescent, Western blotting, immunoprecipitation] Antibody Dilution Company Cat# Mouse antinephrin (G-8) IF: 1:100 Santa Cruz sc-376522 Mouse antinephrin conjugated to agarose IP: 2 mg/400 ml lysate Santa Cruz sc-376522 AC Normal mouse IgG conjugated to agarose IP: 2 mg/400 ml lysate Santa Cruz sc-2343 Rabbit antinephrin WB: 1:1000 Invitrogen PA5-91907 Sheep anti-DCTN1 WB: 1:1000 IF: 1:100 R&D Systems AF6657 Mouse anti-Dynll1 IF: 1:100 Santa Cruz sc-136287 Rabbit anti-Dynll1 WB: 1:1000 IF: 1:100 Thermo Fisher PA5-97920 Rabbit anti-INF2 IF: 1:100 Bethyl Lab A303-427A Rabbit anti-HDAC6 WB: 1:1000 Novus NBP1-78981 Donkey anti-mouse (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32766 Donkey anti-rabbit (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32790 Donkey anti-mouse (Alexa Fluor 594) IF: 1:200 Thermo Fisher A32744 Donkey anti-sheep (Alexa Fluor 488) IF: 1:200 Thermo Fisher A-11015 Mouse anti-rabbit IgG-HRP WB: 1:5000 Santa Cruz sc-2357 Rabbit anti-sheep IgG-HRP WB: 1:5000 Thermo Fisher 31480 Biotin mouse anti-ubiquitin WB: 1:1000 Invitrogen 13-6078-82 Mouse anti b-actin-HRP WB: 1:5000 Santa Cruz sc-47778 Mouse anti-WT1 IF: 1:100 Novus NBP24460700 Mouse anti-acetylated a-tubulin IF: 1:200 Santa Cruz sc-23950 Acti-stain 488 phalloidin Cytoskeleton PHDG1-A ViaFluor microtubule stains (488) Biotium 70062 IF, immunofluorescent; IP, immunoprecipitation; WB, Western blotting; INF2, inverted formin 2; HDAC6, histone deacetylase 6; HRP, horseradish peroxidase. .. 164 KIDNEY360 D ow nladed fom http:urnals.lw w .om /kidey360 by B hD M feP H K a1zE um tQ fN a+ kLhE Z gsIH 4X M ihC yw C X 1A W nY Q p/IlQ rH D 3i3D 0O dR yi7T vS F 4C 3V C 1ybggQ Z X gG 2M w lZ LeI= on 018/2024 Co-Immunoprecipitation and Immunoblotting Cultured podocytes were lysed on ice for 1 hour in Nonidet P40 lysis buffer with 0.5% deoxycholate, cOmplete, Mini Protease Inhibitor Cocktail (Roche, #11836153001), and PhosSTOP phosphatase inhibitor cocktail (Sigma, #4906845001).
Histone Deacetylase Assay:Article Title: Dynein-Mediated Trafficking: A New Mechanism of Diabetic Podocytopathy
Article Snippet: Table 5 Plasmids Company Catalog No. pCDNA3 mouse nephrin Gift from Dr. Lawrence B. Holzman GFP-Rab7 Addgene 12605 GFP-Rab11 Addgene 12674 pEGFP-DCTN1 Addgene 36154 pCMV3-C-GFPSpark-Dynll1 SinoBiochemical MG51246-ACG EB3-tdTomato Addgene 50708 Open in a separate window Plasmids. .. Table 2 Antibody Dilution Company Cat# Mouse antinephrin (G-8) IF: 1:100 Santa Cruz sc-376522 Mouse antinephrin conjugated to agarose IP: 2 μg/400 μl lysate Santa Cruz sc-376522 AC Normal mouse IgG conjugated to agarose IP: 2 μg/400 μl lysate Santa Cruz sc-2343 Rabbit antinephrin WB: 1:1000 Invitrogen PA5-91907 Sheep anti-DCTN1 WB: 1:1000 IF: 1:100 R&D Systems AF6657 Mouse anti-Dynll1 IF: 1:100 Santa Cruz sc-136287 Rabbit anti-Dynll1 WB: 1:1000 IF: 1:100 Thermo Fisher PA5-97920 Rabbit anti-INF2 IF: 1:100 Bethyl Lab A303-427A Rabbit anti-HDAC6 WB: 1:1000 Novus NBP1-78981 Donkey anti-mouse (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32766 Donkey anti-rabbit (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32790 Donkey anti-mouse (Alexa Fluor 594) IF: 1:200 Thermo Fisher A32744 Donkey anti-sheep (Alexa Fluor 488) IF: 1:200 Thermo Fisher A-11015 Mouse anti-rabbit IgG-HRP WB: 1:5000 Santa Cruz sc-2357 Rabbit anti-sheep IgG-HRP WB: 1:5000 Thermo Fisher 31480 Biotin mouse anti-ubiquitin WB: 1:1000 Invitrogen 13-6078-82 Mouse anti β-actin-HRP WB: 1:5000 Santa Cruz sc-47778 Mouse anti-WT1 IF: 1:100 Novus NBP24460700 Mouse anti-acetylated α-tubulin IF: 1:200 Santa Cruz sc-23950 Acti-stain 488 phalloidin Cytoskeleton PHDG1-A ViaFluor microtubule stains (488) Biotium 70062 Open in a separate window IF, immunofluorescent; IP, immunoprecipitation; WB, Western blotting; INF2, inverted formin 2; HDAC6, histone deacetylase 6; HRP, horseradish peroxidase. ..
Article Title: Dynein-Mediated Trafficking: A New Mechanism of Diabetic Podocytopathy
Article Snippet: .. Antibodies and fluorescent probes (immunofluorescent, Western blotting, immunoprecipitation] Antibody Dilution Company Cat# Mouse antinephrin (G-8) IF: 1:100 Santa Cruz sc-376522 Mouse antinephrin conjugated to agarose IP: 2 mg/400 ml lysate Santa Cruz sc-376522 AC Normal mouse IgG conjugated to agarose IP: 2 mg/400 ml lysate Santa Cruz sc-2343 Rabbit antinephrin WB: 1:1000 Invitrogen PA5-91907 Sheep anti-DCTN1 WB: 1:1000 IF: 1:100 R&D Systems AF6657 Mouse anti-Dynll1 IF: 1:100 Santa Cruz sc-136287 Rabbit anti-Dynll1 WB: 1:1000 IF: 1:100 Thermo Fisher PA5-97920 Rabbit anti-INF2 IF: 1:100 Bethyl Lab A303-427A Rabbit anti-HDAC6 WB: 1:1000 Novus NBP1-78981 Donkey anti-mouse (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32766 Donkey anti-rabbit (Alexa Fluor 488) IF: 1:200 Thermo Fisher A32790 Donkey anti-mouse (Alexa Fluor 594) IF: 1:200 Thermo Fisher A32744 Donkey anti-sheep (Alexa Fluor 488) IF: 1:200 Thermo Fisher A-11015 Mouse anti-rabbit IgG-HRP WB: 1:5000 Santa Cruz sc-2357 Rabbit anti-sheep IgG-HRP WB: 1:5000 Thermo Fisher 31480 Biotin mouse anti-ubiquitin WB: 1:1000 Invitrogen 13-6078-82 Mouse anti b-actin-HRP WB: 1:5000 Santa Cruz sc-47778 Mouse anti-WT1 IF: 1:100 Novus NBP24460700 Mouse anti-acetylated a-tubulin IF: 1:200 Santa Cruz sc-23950 Acti-stain 488 phalloidin Cytoskeleton PHDG1-A ViaFluor microtubule stains (488) Biotium 70062 IF, immunofluorescent; IP, immunoprecipitation; WB, Western blotting; INF2, inverted formin 2; HDAC6, histone deacetylase 6; HRP, horseradish peroxidase. .. 164 KIDNEY360 D ow nladed fom http:urnals.lw w .om /kidey360 by B hD M feP H K a1zE um tQ fN a+ kLhE Z gsIH 4X M ihC yw C X 1A W nY Q p/IlQ rH D 3i3D 0O dR yi7T vS F 4C 3V C 1ybggQ Z X gG 2M w lZ LeI= on 018/2024 Co-Immunoprecipitation and Immunoblotting Cultured podocytes were lysed on ice for 1 hour in Nonidet P40 lysis buffer with 0.5% deoxycholate, cOmplete, Mini Protease Inhibitor Cocktail (Roche, #11836153001), and PhosSTOP phosphatase inhibitor cocktail (Sigma, #4906845001).
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