Immunofluorescence:Article Title: MAGIs regulate aPKC to enable balanced distribution of intercellular tension for epithelial sheet homeostasis
Article Snippet: .. The target sequences were as follows; Mouse Afadin: 5′- GAGGAGAGCATGCGCATGTC-3′ Mouse ASPP1: 5′-GACTACAGCAAGATCATGAA-3′ Mouse ASPP2: 5′-GAGCAAGGAACCATCAGGCA-3′ Mouse MAGI-1: 5′-GGGGACCCCAGGGCGAGCTG-3′ Mouse MAGI-3: 5′-GAGGACCTGATCAACTGGAT-3′ The following primary antibodies were used for immunofluorescence (IF) microscopy, immunoprecipitation (IP) and immunoblotting (IB): rat anti-ZO-1 (DSHB; 1:50, IF; 1:500, IB); rabbit anti-ZO-2 (Zymed; 1:1000 IB); mouse anti-AF6 (afadin; BD Biosciences; 1:200, IF); rabbit anti-Par-3 (Merck Millipore; 1:300, IF); mouse anti-MAGI-1 (ss-5; 1:100, IF; 1:500, IB), mouse anti-MAGI-3 (46; 1:100, IF; 1:500, IB) and rabbit anti-PKCζ antibodies (Santa Cruz Biotechnology; 1:100, IF); mouse anti-phospho-myosin light chain 2 (Ser19; used in Supplementary Figs. and ; 1:100, IF), rabbit anti-phospho-myosin light chain 2 (Ser19; used in Fig. ; 1:100, IF) and rabbit anti-Ezrin (CST; 1:100, IF); rabbit anti-ROCK1 (abcam; 1:200, IF); rabbit anti-Claudin-3 (Life Technologies; 1:100, IF); mouse anti-GFP (1:100, IF; 1:1000, IB) and rat anti-HA (Roche; 1:1000 IB); mouse anti-DYKDDDDK (FLAG; 1:1000 IB) and rat anti-E-cadherin (ECCD-2; Wako Pure Chemicals; 1:100, IF). ..
Article Title: MARCKS-related protein regulates cytoskeletal organization at cell–cell and cell–substrate contacts in epithelial cells
Article Snippet: Image preparation was performed with ImageJ and assembly was carried out in Adobe Photoshop. .. Rat anti-ZO-1 [40.76, used at 1:500 for immunoblots (IB) and 1:100 for immunofluorescence, IF] has been previously described ( Anderson et al., 1988 ), rabbit anti-occludin polyclonal antibody was from ProteinTech (13409-1-AP; 1:1000 IB, 1:800 IF), rabbit anti-ZO-2 (38-9100; 1:1000 IB, 1:100, IF) and anti-phospho-FAK(Tyr397) (700255; 1:1000 IB) antibodies were from ThermoFisher; mouse anti-FAK (610087; 1:1000 IB, 1:100 IF) and mouse anti-E-cadherin (61081)(1:1000 IB, 1:100 IF) antibodies were from BD Transduction Laboratories, rabbit anti-phospho-MLC(S19) (3671P; 1:100 IF), -Src (2109; 1:1000 IB), -phospho-Src(Tyr416) (2101;1:1000 IB) and mouse anti-Myc tag (9B11; 1:100 IF) antibodies were from Cell Signaling Technology; rat anti-E-cadherin (U3254; 1:200 IF), mouse anti-α-actinin (1:100 IF), mouse anti-γ-tubulin (1:1000 IB) and mouse anti-vinculin (V9131; 1:1000 IB, 1:100 IF) were from were from Sigma-Aldrich; rabbit anti-myosin 2B (909901) and 2A (909801) (1:1000 IF, 1:250 IB) antibodies were from BioLegend; rabbit anti-GFP antibody (Ab290); 1:2000 IB) was from Abcam; mouse anti-actin antibody (MAB1501R; 1:2000 IF) was from Millipore; mouse monoclonal antibody to GP135/podocalyxin (1:100 IF) was kindly provided by George Ojakian, State University of New York, NY. ..
Microscopy:Article Title: MAGIs regulate aPKC to enable balanced distribution of intercellular tension for epithelial sheet homeostasis
Article Snippet: .. The target sequences were as follows; Mouse Afadin: 5′- GAGGAGAGCATGCGCATGTC-3′ Mouse ASPP1: 5′-GACTACAGCAAGATCATGAA-3′ Mouse ASPP2: 5′-GAGCAAGGAACCATCAGGCA-3′ Mouse MAGI-1: 5′-GGGGACCCCAGGGCGAGCTG-3′ Mouse MAGI-3: 5′-GAGGACCTGATCAACTGGAT-3′ The following primary antibodies were used for immunofluorescence (IF) microscopy, immunoprecipitation (IP) and immunoblotting (IB): rat anti-ZO-1 (DSHB; 1:50, IF; 1:500, IB); rabbit anti-ZO-2 (Zymed; 1:1000 IB); mouse anti-AF6 (afadin; BD Biosciences; 1:200, IF); rabbit anti-Par-3 (Merck Millipore; 1:300, IF); mouse anti-MAGI-1 (ss-5; 1:100, IF; 1:500, IB), mouse anti-MAGI-3 (46; 1:100, IF; 1:500, IB) and rabbit anti-PKCζ antibodies (Santa Cruz Biotechnology; 1:100, IF); mouse anti-phospho-myosin light chain 2 (Ser19; used in Supplementary Figs. and ; 1:100, IF), rabbit anti-phospho-myosin light chain 2 (Ser19; used in Fig. ; 1:100, IF) and rabbit anti-Ezrin (CST; 1:100, IF); rabbit anti-ROCK1 (abcam; 1:200, IF); rabbit anti-Claudin-3 (Life Technologies; 1:100, IF); mouse anti-GFP (1:100, IF; 1:1000, IB) and rat anti-HA (Roche; 1:1000 IB); mouse anti-DYKDDDDK (FLAG; 1:1000 IB) and rat anti-E-cadherin (ECCD-2; Wako Pure Chemicals; 1:100, IF). ..
Article Title: Different effects of ZO-1, ZO-2 and ZO-3 silencing on kidney collecting duct principal cell proliferation and adhesion
Article Snippet: .. The following antibodies were used for Western blot and microscopy analysis: rat anti-ZO-1 (Millipore, NG1853799), rabbit anti-ZO-1 (Invitrogen, 61–7300), rabbit anti-ZO-2 (Invitrogen, 71–1400), rabbit anti-ZO-3 (Invitrogen, 36–4100), rabbit anti-ZONAB (Novex, 40–2800), rabbit anti-CycD1 (Santa Cruz, sc-717), mouse anti-CycD1 (Santa Cruz, sc-450), mouse anti-PCNA (Santa Cruz, sc-56), rabbit anti-p21 (Santa Cruz, sc-397), rabbit anti-calreticulin (Cell Signaling, 12238), rabbit anti-HDAC3 (Cell Signaling, 2632), mouse anti-α-tubulin (Sigma-Aldrich, T9026), mouse anti-β-actin (Sigma-Aldrich, A5441), mouse anti-claudin-4 (Invitrogen, 32–9400), rabbit claudin-8 (Invitrogen, 40–0700Z), mouse anti-occludin (Invitrogen, 33–1500), rabbit anti-β1-integrin (Abcam, 52971) and rat anti-β1-integrin (9EG7) (BD Biosciences, 553715). .. Secondary antibodies used were as follows: Horseradish peroxidase (HRP)-conjugated goat anti-mouse (BD, 554002), HRP-conjugated goat anti-rabbit (BD, 554021), HRP-conjugated goat anti-rat (Invitrogen, 629520), goat anti-mouse Alexa Fluor 488 (Invitrogen, A-11017), goat anti-rabbit Alexa Fluor 488 (Invitrogen, A-11070), goat anti-rat Alexa Fluor 488 (Jackson ImmunoResearch Laboratories, 112-545-003) goat anti-rabbit CY3 (Jackson ImmunoResearch Laboratories, 111-165-003), goat anti-mouse CY3 (Jackson ImmunoResearch Laboratories, 115-166-003), goat anti-rat Alexa Fluor 546 (Invitrogen, {"type":"entrez-nucleotide","attrs":{"text":"A11081","term_id":"489258","term_text":"A11081"}} A11081 ).
Article Title: Different effects of ZO-1, ZO-2 and ZO-3 silencing on kidney collecting duct principal cell proliferation and adhesion
Article Snippet: .. Antibodies and reagents The following antibodies were used for Western blot and microscopy analysis: rat anti-ZO-1 (Millipore, NG1853799), rabbit anti-ZO-1 (Invitrogen, 61–7300), rabbit anti-ZO-2 (Invitrogen, 71–1400), rabbit anti-ZO-3 (Invitrogen, 36–4100), rabbit anti-ZONAB (Novex, 40–2800), rabbit anti-CycD1 (Santa Cruz, sc-717), mouse anti-CycD1 (Santa Cruz, sc-450), mouse anti-PCNA (Santa Cruz, sc-56), rabbit anti-p21 (Santa Cruz, sc-397), rabbit anti-calreticulin (Cell Signaling, 12238), rabbit anti-HDAC3 (Cell Signaling, 2632), mouse anti-α-tubulin (Sigma-Aldrich, T9026), mouse anti-β-actin (Sigma-Aldrich, A5441), mouse anti-claudin-4 (Invitrogen, 32–9400), rabbit claudin-8 (Invitrogen, 40–0700Z), mouse anti-occludin (Invitrogen, 33–1500), rabbit anti-β1-integrin (Abcam, 52971) and rat anti-β1-integrin (9EG7) (BD Biosciences, 553715). .. Secondary antibodies used were as follows: Horseradish peroxidase (HRP)-conjugated goat anti-mouse (BD, 554002), HRP-conjugated goat anti-rabbit (BD, 554021), HRP-conjugated goat anti-rat (Invitrogen, 629520), goat anti-mouse Alexa Fluor 488 (Invitrogen, A-11017), goat anti-rabbit Alexa Fluor 488 (Invitrogen, A-11070), goat anti-rat Alexa Fluor 488 (Jackson ImmunoResearch Laboratories, 112-545-003) goat anti-rabbit CY3 (Jackson ImmunoResearch Laboratories, 111-165-003), goat anti-mouse CY3 (Jackson ImmunoResearch Laboratories, 115-166-003), goat anti-rat Alexa Fluor 546 (Invitrogen, {"type":"entrez-nucleotide","attrs":{"text":"A11081","term_id":"489258","term_text":"A11081"}} A11081 ).
Immunoprecipitation:Article Title: MAGIs regulate aPKC to enable balanced distribution of intercellular tension for epithelial sheet homeostasis
Article Snippet: .. The target sequences were as follows; Mouse Afadin: 5′- GAGGAGAGCATGCGCATGTC-3′ Mouse ASPP1: 5′-GACTACAGCAAGATCATGAA-3′ Mouse ASPP2: 5′-GAGCAAGGAACCATCAGGCA-3′ Mouse MAGI-1: 5′-GGGGACCCCAGGGCGAGCTG-3′ Mouse MAGI-3: 5′-GAGGACCTGATCAACTGGAT-3′ The following primary antibodies were used for immunofluorescence (IF) microscopy, immunoprecipitation (IP) and immunoblotting (IB): rat anti-ZO-1 (DSHB; 1:50, IF; 1:500, IB); rabbit anti-ZO-2 (Zymed; 1:1000 IB); mouse anti-AF6 (afadin; BD Biosciences; 1:200, IF); rabbit anti-Par-3 (Merck Millipore; 1:300, IF); mouse anti-MAGI-1 (ss-5; 1:100, IF; 1:500, IB), mouse anti-MAGI-3 (46; 1:100, IF; 1:500, IB) and rabbit anti-PKCζ antibodies (Santa Cruz Biotechnology; 1:100, IF); mouse anti-phospho-myosin light chain 2 (Ser19; used in Supplementary Figs. and ; 1:100, IF), rabbit anti-phospho-myosin light chain 2 (Ser19; used in Fig. ; 1:100, IF) and rabbit anti-Ezrin (CST; 1:100, IF); rabbit anti-ROCK1 (abcam; 1:200, IF); rabbit anti-Claudin-3 (Life Technologies; 1:100, IF); mouse anti-GFP (1:100, IF; 1:1000, IB) and rat anti-HA (Roche; 1:1000 IB); mouse anti-DYKDDDDK (FLAG; 1:1000 IB) and rat anti-E-cadherin (ECCD-2; Wako Pure Chemicals; 1:100, IF). ..
Western Blot:Article Title: MAGIs regulate aPKC to enable balanced distribution of intercellular tension for epithelial sheet homeostasis
Article Snippet: .. The target sequences were as follows; Mouse Afadin: 5′- GAGGAGAGCATGCGCATGTC-3′ Mouse ASPP1: 5′-GACTACAGCAAGATCATGAA-3′ Mouse ASPP2: 5′-GAGCAAGGAACCATCAGGCA-3′ Mouse MAGI-1: 5′-GGGGACCCCAGGGCGAGCTG-3′ Mouse MAGI-3: 5′-GAGGACCTGATCAACTGGAT-3′ The following primary antibodies were used for immunofluorescence (IF) microscopy, immunoprecipitation (IP) and immunoblotting (IB): rat anti-ZO-1 (DSHB; 1:50, IF; 1:500, IB); rabbit anti-ZO-2 (Zymed; 1:1000 IB); mouse anti-AF6 (afadin; BD Biosciences; 1:200, IF); rabbit anti-Par-3 (Merck Millipore; 1:300, IF); mouse anti-MAGI-1 (ss-5; 1:100, IF; 1:500, IB), mouse anti-MAGI-3 (46; 1:100, IF; 1:500, IB) and rabbit anti-PKCζ antibodies (Santa Cruz Biotechnology; 1:100, IF); mouse anti-phospho-myosin light chain 2 (Ser19; used in Supplementary Figs. and ; 1:100, IF), rabbit anti-phospho-myosin light chain 2 (Ser19; used in Fig. ; 1:100, IF) and rabbit anti-Ezrin (CST; 1:100, IF); rabbit anti-ROCK1 (abcam; 1:200, IF); rabbit anti-Claudin-3 (Life Technologies; 1:100, IF); mouse anti-GFP (1:100, IF; 1:1000, IB) and rat anti-HA (Roche; 1:1000 IB); mouse anti-DYKDDDDK (FLAG; 1:1000 IB) and rat anti-E-cadherin (ECCD-2; Wako Pure Chemicals; 1:100, IF). ..
Article Title: Different effects of ZO-1, ZO-2 and ZO-3 silencing on kidney collecting duct principal cell proliferation and adhesion
Article Snippet: .. The following antibodies were used for Western blot and microscopy analysis: rat anti-ZO-1 (Millipore, NG1853799), rabbit anti-ZO-1 (Invitrogen, 61–7300), rabbit anti-ZO-2 (Invitrogen, 71–1400), rabbit anti-ZO-3 (Invitrogen, 36–4100), rabbit anti-ZONAB (Novex, 40–2800), rabbit anti-CycD1 (Santa Cruz, sc-717), mouse anti-CycD1 (Santa Cruz, sc-450), mouse anti-PCNA (Santa Cruz, sc-56), rabbit anti-p21 (Santa Cruz, sc-397), rabbit anti-calreticulin (Cell Signaling, 12238), rabbit anti-HDAC3 (Cell Signaling, 2632), mouse anti-α-tubulin (Sigma-Aldrich, T9026), mouse anti-β-actin (Sigma-Aldrich, A5441), mouse anti-claudin-4 (Invitrogen, 32–9400), rabbit claudin-8 (Invitrogen, 40–0700Z), mouse anti-occludin (Invitrogen, 33–1500), rabbit anti-β1-integrin (Abcam, 52971) and rat anti-β1-integrin (9EG7) (BD Biosciences, 553715). .. Secondary antibodies used were as follows: Horseradish peroxidase (HRP)-conjugated goat anti-mouse (BD, 554002), HRP-conjugated goat anti-rabbit (BD, 554021), HRP-conjugated goat anti-rat (Invitrogen, 629520), goat anti-mouse Alexa Fluor 488 (Invitrogen, A-11017), goat anti-rabbit Alexa Fluor 488 (Invitrogen, A-11070), goat anti-rat Alexa Fluor 488 (Jackson ImmunoResearch Laboratories, 112-545-003) goat anti-rabbit CY3 (Jackson ImmunoResearch Laboratories, 111-165-003), goat anti-mouse CY3 (Jackson ImmunoResearch Laboratories, 115-166-003), goat anti-rat Alexa Fluor 546 (Invitrogen, {"type":"entrez-nucleotide","attrs":{"text":"A11081","term_id":"489258","term_text":"A11081"}} A11081 ).
Article Title: MARCKS-related protein regulates cytoskeletal organization at cell–cell and cell–substrate contacts in epithelial cells
Article Snippet: Image preparation was performed with ImageJ and assembly was carried out in Adobe Photoshop. .. Rat anti-ZO-1 [40.76, used at 1:500 for immunoblots (IB) and 1:100 for immunofluorescence, IF] has been previously described ( Anderson et al., 1988 ), rabbit anti-occludin polyclonal antibody was from ProteinTech (13409-1-AP; 1:1000 IB, 1:800 IF), rabbit anti-ZO-2 (38-9100; 1:1000 IB, 1:100, IF) and anti-phospho-FAK(Tyr397) (700255; 1:1000 IB) antibodies were from ThermoFisher; mouse anti-FAK (610087; 1:1000 IB, 1:100 IF) and mouse anti-E-cadherin (61081)(1:1000 IB, 1:100 IF) antibodies were from BD Transduction Laboratories, rabbit anti-phospho-MLC(S19) (3671P; 1:100 IF), -Src (2109; 1:1000 IB), -phospho-Src(Tyr416) (2101;1:1000 IB) and mouse anti-Myc tag (9B11; 1:100 IF) antibodies were from Cell Signaling Technology; rat anti-E-cadherin (U3254; 1:200 IF), mouse anti-α-actinin (1:100 IF), mouse anti-γ-tubulin (1:1000 IB) and mouse anti-vinculin (V9131; 1:1000 IB, 1:100 IF) were from were from Sigma-Aldrich; rabbit anti-myosin 2B (909901) and 2A (909801) (1:1000 IF, 1:250 IB) antibodies were from BioLegend; rabbit anti-GFP antibody (Ab290); 1:2000 IB) was from Abcam; mouse anti-actin antibody (MAB1501R; 1:2000 IF) was from Millipore; mouse monoclonal antibody to GP135/podocalyxin (1:100 IF) was kindly provided by George Ojakian, State University of New York, NY. ..
Article Title: Different effects of ZO-1, ZO-2 and ZO-3 silencing on kidney collecting duct principal cell proliferation and adhesion
Article Snippet: .. Antibodies and reagents The following antibodies were used for Western blot and microscopy analysis: rat anti-ZO-1 (Millipore, NG1853799), rabbit anti-ZO-1 (Invitrogen, 61–7300), rabbit anti-ZO-2 (Invitrogen, 71–1400), rabbit anti-ZO-3 (Invitrogen, 36–4100), rabbit anti-ZONAB (Novex, 40–2800), rabbit anti-CycD1 (Santa Cruz, sc-717), mouse anti-CycD1 (Santa Cruz, sc-450), mouse anti-PCNA (Santa Cruz, sc-56), rabbit anti-p21 (Santa Cruz, sc-397), rabbit anti-calreticulin (Cell Signaling, 12238), rabbit anti-HDAC3 (Cell Signaling, 2632), mouse anti-α-tubulin (Sigma-Aldrich, T9026), mouse anti-β-actin (Sigma-Aldrich, A5441), mouse anti-claudin-4 (Invitrogen, 32–9400), rabbit claudin-8 (Invitrogen, 40–0700Z), mouse anti-occludin (Invitrogen, 33–1500), rabbit anti-β1-integrin (Abcam, 52971) and rat anti-β1-integrin (9EG7) (BD Biosciences, 553715). .. Secondary antibodies used were as follows: Horseradish peroxidase (HRP)-conjugated goat anti-mouse (BD, 554002), HRP-conjugated goat anti-rabbit (BD, 554021), HRP-conjugated goat anti-rat (Invitrogen, 629520), goat anti-mouse Alexa Fluor 488 (Invitrogen, A-11017), goat anti-rabbit Alexa Fluor 488 (Invitrogen, A-11070), goat anti-rat Alexa Fluor 488 (Jackson ImmunoResearch Laboratories, 112-545-003) goat anti-rabbit CY3 (Jackson ImmunoResearch Laboratories, 111-165-003), goat anti-mouse CY3 (Jackson ImmunoResearch Laboratories, 115-166-003), goat anti-rat Alexa Fluor 546 (Invitrogen, {"type":"entrez-nucleotide","attrs":{"text":"A11081","term_id":"489258","term_text":"A11081"}} A11081 ).
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