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  • 99
    Millipore β actin
    CCP6 or CCP1 deficiency promotes cell reprogramming. a Ccp1 - or Ccp6 - deficient MEFs were lyzed for immunoblotting. <t>β-actin</t> was used as a loading control. b WT or CCPs-deficient MEFs were infected by OSKM factors containing retrovirus and cultured in ESC media for 3 weeks. Alkaline phosphatase (AP)-positive colony numbers per 10 4 cells were calculated and shown as means ± S.D. **, P
    β Actin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 67284 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Santa Cruz Biotechnology β actin
    Activation of the AKT- IκB kinase (IKK)-inhibitors of NF-κB (IκB) pathway is associated with the CK2 down-regulation-mediated nuclear import of NF-κB. ( A ) MCF-7 and HCT116 cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days. The level of each protein was determined by immunoblot analysis using specific antibodies ( left ). Representative data from three independent experiments are shown. <t>β-Actin</t> was used as a control. Graphs represent the quantitation of p-AKT, IKK, and IκBα relative to β-actin and that of p-IKK and p-IκBα relative to the unphosphorylated proteins ( right ). ( B ) Cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days in the absence or presence of 1 μM triciribine (TCN). Cytoplasm and nuclei were isolated from cells, and both extracts were visualized by immunoblotting. α-Tubulin (cytoplasmic marker) and histone H3 (nuclear marker) were quantified as loading controls ( left ). Representative data from three independent experiments are shown. Graphs represent the quantitation of RelA/p65 relative to the subcellular markers ( right ). exp, exposure. Data are mean ± SEM. * p
    β Actin, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 95/100, based on 50832 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Cell Signaling Technology Inc β actin
    Western blot representative of omentin in diabetic animals. (A) in tissues: Molecular Weight Marker (M); Mesenteric Adipose Tissue (MES), Retroperitoneal (RET), Epididimal (EPI), Brown Adipose Tissue (BAT), Liver, and Serum. For this result, 3 independent replicates were performed. (B) Representative membrane with 4 sample of omentin in muscle (one per group) compared to MES, showed that omentin was not found in muscle (I = 6 sample per group). (C) Serum Omentin Values (ELISA) ( n = 10 sample per group). (D) Omentin (40 kDa) in MES in the experimental protocols. C, control group; RT, resistance training; AT, aerobic training; CT, combined training. Results are presented as the relative density after normalizing with <t>β-actin</t> protein. Data are expressed as means ± SD ( n = 6 sample per group). P
    β Actin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 38293 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore monoclonal anti beta actin antibody
    Western blot representative of omentin in diabetic animals. (A) in tissues: Molecular Weight Marker (M); Mesenteric Adipose Tissue (MES), Retroperitoneal (RET), Epididimal (EPI), Brown Adipose Tissue (BAT), Liver, and Serum. For this result, 3 independent replicates were performed. (B) Representative membrane with 4 sample of omentin in muscle (one per group) compared to MES, showed that omentin was not found in muscle (I = 6 sample per group). (C) Serum Omentin Values (ELISA) ( n = 10 sample per group). (D) Omentin (40 kDa) in MES in the experimental protocols. C, control group; RT, resistance training; AT, aerobic training; CT, combined training. Results are presented as the relative density after normalizing with <t>β-actin</t> protein. Data are expressed as means ± SD ( n = 6 sample per group). P
    Monoclonal Anti Beta Actin Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 33074 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore anti β actin
    Pellino2 ubiquitinates IRAK1 and suppress the inhibitory effects of IRAK1 on NLRP3. a Immunoblot analysis of myc and IRAK1 in lysates (Input) and immunoprecipitated (IP) myc samples from HEK293T cells transfected with myc-tagged Pellino2 and untagged IRAK1. b Immunoblot analysis of HA, IRAK1 and FLAG in lysates (Input) and IP IRAK1 samples from HEK293T cells transfected with FLAG-tagged Pellino2, FLAG-tagged Pellino2 RING mutant, untagged IRAK1, and HA-Ubiquitin. c, d Immunoblot analysis of Ubiquitin (short exposure (SE) or long exposure (LE)) and IRAK1 in lysates (Input) and IP IRAK1 samples or IRAK1 and ubq in TUBE-Ubq elution and cell lysates ( d ) from WT and Peli2 −/− BMDMs treated with 100 ng/ml of LPS for the indicated times. e Immunoblot analysis of NLRP3 and IRAK1 in lysates (Input) and IP IRAK1 samples from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. f ELISA of IL-1β (left panel) and TNF (right panel) in medium from primary WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for 3 h and then with 2.5 mM ATP or 5 mM Nigericin for 1 h. UT, untreated. g Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates of WT and Irak1 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h and 2.5 mM ATP for 1 h. h Immunoblot analysis of HA, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from HEK293T cells transfected with V5-tagged NLRP3, untagged IRAK1, untagged kinase dead IRAK1 (IRAK1-KD), and HA-ubiquitin. <t>β-Actin</t> was used as loading controls. i Immunoblot analysis of Ubiquitin, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from immortalized WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. j Immortalized WT and Irak1 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING) or Pellino2 FHA mutant (FHA). Immunoblot analysis of myc and NLRP3 in lysates (Input) and IP myc samples from virus-infected BMDMs treated with 100 ng/ml LPS for indicated times. * p
    Anti β Actin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 26841 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam β actin
    Pellino2 ubiquitinates IRAK1 and suppress the inhibitory effects of IRAK1 on NLRP3. a Immunoblot analysis of myc and IRAK1 in lysates (Input) and immunoprecipitated (IP) myc samples from HEK293T cells transfected with myc-tagged Pellino2 and untagged IRAK1. b Immunoblot analysis of HA, IRAK1 and FLAG in lysates (Input) and IP IRAK1 samples from HEK293T cells transfected with FLAG-tagged Pellino2, FLAG-tagged Pellino2 RING mutant, untagged IRAK1, and HA-Ubiquitin. c, d Immunoblot analysis of Ubiquitin (short exposure (SE) or long exposure (LE)) and IRAK1 in lysates (Input) and IP IRAK1 samples or IRAK1 and ubq in TUBE-Ubq elution and cell lysates ( d ) from WT and Peli2 −/− BMDMs treated with 100 ng/ml of LPS for the indicated times. e Immunoblot analysis of NLRP3 and IRAK1 in lysates (Input) and IP IRAK1 samples from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. f ELISA of IL-1β (left panel) and TNF (right panel) in medium from primary WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for 3 h and then with 2.5 mM ATP or 5 mM Nigericin for 1 h. UT, untreated. g Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates of WT and Irak1 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h and 2.5 mM ATP for 1 h. h Immunoblot analysis of HA, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from HEK293T cells transfected with V5-tagged NLRP3, untagged IRAK1, untagged kinase dead IRAK1 (IRAK1-KD), and HA-ubiquitin. <t>β-Actin</t> was used as loading controls. i Immunoblot analysis of Ubiquitin, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from immortalized WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. j Immortalized WT and Irak1 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING) or Pellino2 FHA mutant (FHA). Immunoblot analysis of myc and NLRP3 in lysates (Input) and IP myc samples from virus-infected BMDMs treated with 100 ng/ml LPS for indicated times. * p
    β Actin, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 25356 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher β mercaptoethanol
    Pellino2 ubiquitinates IRAK1 and suppress the inhibitory effects of IRAK1 on NLRP3. a Immunoblot analysis of myc and IRAK1 in lysates (Input) and immunoprecipitated (IP) myc samples from HEK293T cells transfected with myc-tagged Pellino2 and untagged IRAK1. b Immunoblot analysis of HA, IRAK1 and FLAG in lysates (Input) and IP IRAK1 samples from HEK293T cells transfected with FLAG-tagged Pellino2, FLAG-tagged Pellino2 RING mutant, untagged IRAK1, and HA-Ubiquitin. c, d Immunoblot analysis of Ubiquitin (short exposure (SE) or long exposure (LE)) and IRAK1 in lysates (Input) and IP IRAK1 samples or IRAK1 and ubq in TUBE-Ubq elution and cell lysates ( d ) from WT and Peli2 −/− BMDMs treated with 100 ng/ml of LPS for the indicated times. e Immunoblot analysis of NLRP3 and IRAK1 in lysates (Input) and IP IRAK1 samples from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. f ELISA of IL-1β (left panel) and TNF (right panel) in medium from primary WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for 3 h and then with 2.5 mM ATP or 5 mM Nigericin for 1 h. UT, untreated. g Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates of WT and Irak1 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h and 2.5 mM ATP for 1 h. h Immunoblot analysis of HA, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from HEK293T cells transfected with V5-tagged NLRP3, untagged IRAK1, untagged kinase dead IRAK1 (IRAK1-KD), and HA-ubiquitin. <t>β-Actin</t> was used as loading controls. i Immunoblot analysis of Ubiquitin, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from immortalized WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. j Immortalized WT and Irak1 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING) or Pellino2 FHA mutant (FHA). Immunoblot analysis of myc and NLRP3 in lysates (Input) and IP myc samples from virus-infected BMDMs treated with 100 ng/ml LPS for indicated times. * p
    β Mercaptoethanol, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 18648 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore β mercaptoethanol
    Pellino2 ubiquitinates IRAK1 and suppress the inhibitory effects of IRAK1 on NLRP3. a Immunoblot analysis of myc and IRAK1 in lysates (Input) and immunoprecipitated (IP) myc samples from HEK293T cells transfected with myc-tagged Pellino2 and untagged IRAK1. b Immunoblot analysis of HA, IRAK1 and FLAG in lysates (Input) and IP IRAK1 samples from HEK293T cells transfected with FLAG-tagged Pellino2, FLAG-tagged Pellino2 RING mutant, untagged IRAK1, and HA-Ubiquitin. c, d Immunoblot analysis of Ubiquitin (short exposure (SE) or long exposure (LE)) and IRAK1 in lysates (Input) and IP IRAK1 samples or IRAK1 and ubq in TUBE-Ubq elution and cell lysates ( d ) from WT and Peli2 −/− BMDMs treated with 100 ng/ml of LPS for the indicated times. e Immunoblot analysis of NLRP3 and IRAK1 in lysates (Input) and IP IRAK1 samples from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. f ELISA of IL-1β (left panel) and TNF (right panel) in medium from primary WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for 3 h and then with 2.5 mM ATP or 5 mM Nigericin for 1 h. UT, untreated. g Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates of WT and Irak1 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h and 2.5 mM ATP for 1 h. h Immunoblot analysis of HA, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from HEK293T cells transfected with V5-tagged NLRP3, untagged IRAK1, untagged kinase dead IRAK1 (IRAK1-KD), and HA-ubiquitin. <t>β-Actin</t> was used as loading controls. i Immunoblot analysis of Ubiquitin, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from immortalized WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. j Immortalized WT and Irak1 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING) or Pellino2 FHA mutant (FHA). Immunoblot analysis of myc and NLRP3 in lysates (Input) and IP myc samples from virus-infected BMDMs treated with 100 ng/ml LPS for indicated times. * p
    β Mercaptoethanol, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 28107 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore β glycerophosphate
    Pellino2 ubiquitinates IRAK1 and suppress the inhibitory effects of IRAK1 on NLRP3. a Immunoblot analysis of myc and IRAK1 in lysates (Input) and immunoprecipitated (IP) myc samples from HEK293T cells transfected with myc-tagged Pellino2 and untagged IRAK1. b Immunoblot analysis of HA, IRAK1 and FLAG in lysates (Input) and IP IRAK1 samples from HEK293T cells transfected with FLAG-tagged Pellino2, FLAG-tagged Pellino2 RING mutant, untagged IRAK1, and HA-Ubiquitin. c, d Immunoblot analysis of Ubiquitin (short exposure (SE) or long exposure (LE)) and IRAK1 in lysates (Input) and IP IRAK1 samples or IRAK1 and ubq in TUBE-Ubq elution and cell lysates ( d ) from WT and Peli2 −/− BMDMs treated with 100 ng/ml of LPS for the indicated times. e Immunoblot analysis of NLRP3 and IRAK1 in lysates (Input) and IP IRAK1 samples from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. f ELISA of IL-1β (left panel) and TNF (right panel) in medium from primary WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for 3 h and then with 2.5 mM ATP or 5 mM Nigericin for 1 h. UT, untreated. g Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates of WT and Irak1 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h and 2.5 mM ATP for 1 h. h Immunoblot analysis of HA, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from HEK293T cells transfected with V5-tagged NLRP3, untagged IRAK1, untagged kinase dead IRAK1 (IRAK1-KD), and HA-ubiquitin. <t>β-Actin</t> was used as loading controls. i Immunoblot analysis of Ubiquitin, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from immortalized WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. j Immortalized WT and Irak1 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING) or Pellino2 FHA mutant (FHA). Immunoblot analysis of myc and NLRP3 in lysates (Input) and IP myc samples from virus-infected BMDMs treated with 100 ng/ml LPS for indicated times. * p
    β Glycerophosphate, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 12918 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Santa Cruz Biotechnology anti β actin
    Staufen1 knockdown inhibits Prostate Cancer cell migration. Migration assays were performed after 48 h of Staufen1 knockdown and a is a representative western blot of Staufen1 expression with <t>β-actin</t> as a loading control in PC3 and DU145 cells. Each representative blot is cropped to show an n = 1 for each cell line from their respective full-length blot. Note that the DU145 cells used for Fig. 4 were also used for Fig. 5 experiments and therefore a different representative image from the same blot was selected for each figure. Quantification is represented normalized to CTL (n = 3). Data are Mean ± SD, One-Sample T-Test, **P
    Anti β Actin, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 14405 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore anti β actin antibody
    Staufen1 knockdown inhibits Prostate Cancer cell migration. Migration assays were performed after 48 h of Staufen1 knockdown and a is a representative western blot of Staufen1 expression with <t>β-actin</t> as a loading control in PC3 and DU145 cells. Each representative blot is cropped to show an n = 1 for each cell line from their respective full-length blot. Note that the DU145 cells used for Fig. 4 were also used for Fig. 5 experiments and therefore a different representative image from the same blot was selected for each figure. Quantification is represented normalized to CTL (n = 3). Data are Mean ± SD, One-Sample T-Test, **P
    Anti β Actin Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 9783 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Cell Signaling Technology Inc anti β actin
    Staufen1 knockdown inhibits Prostate Cancer cell migration. Migration assays were performed after 48 h of Staufen1 knockdown and a is a representative western blot of Staufen1 expression with <t>β-actin</t> as a loading control in PC3 and DU145 cells. Each representative blot is cropped to show an n = 1 for each cell line from their respective full-length blot. Note that the DU145 cells used for Fig. 4 were also used for Fig. 5 experiments and therefore a different representative image from the same blot was selected for each figure. Quantification is represented normalized to CTL (n = 3). Data are Mean ± SD, One-Sample T-Test, **P
    Anti β Actin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 9551 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    R&D Systems tgf β1
    Monocyte chemoattractant protein‐1 mediates tumor growth factor <t>β1‐primed</t> fibrogenic activation of hepatic stellate cells. (A) Experimental approach to analyze the fibrogenic activation of hSCs. LX‐2 cells were treated with <t>TGF‐ß1</t> for 1 hour and then supplied with fresh medium. MCP‐1 concentration in the culture supernatant was assessed at the indicated times. During cell cultivation, the culture medium was changed daily with or without the presence of MCP‐1 blocking antibody. At the end of the culturing phase, the cell count and type IV collagen protein levels were analyzed. (B) Assessment of extracellular secretion of MCP‐1 in the culture supernatant. Data represent mean ± SD. (C) Cell growth at 120 hours after transient TGF‐β1 stimulation (0, 10 ng/mL and 100 ng/mL) in the presence of either IgG control antibody or MCP‐1 blocking antibody ( # P
    Tgf β1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 99/100, based on 8820 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc β catenin
    Liver, 22-month-old male C57BL/6J mouse. Hepatoblastoma (left) and adjacent hepatocellular adenoma (right). <t>β-catenin</t> immunohistochemistry reveals a strong nucleocytoplasmic signal in hepatoblastoma tumor cells, while neoplastic hepatocytes in the surrounding adenoma display a delicate membranous staining pattern. β-catenin immunohistochemistry (IHC); hematoxylin counterstain. Scale bar: 100 µm.
    β Catenin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 8344 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore mouse anti β actin
    Liver, 22-month-old male C57BL/6J mouse. Hepatoblastoma (left) and adjacent hepatocellular adenoma (right). <t>β-catenin</t> immunohistochemistry reveals a strong nucleocytoplasmic signal in hepatoblastoma tumor cells, while neoplastic hepatocytes in the surrounding adenoma display a delicate membranous staining pattern. β-catenin immunohistochemistry (IHC); hematoxylin counterstain. Scale bar: 100 µm.
    Mouse Anti β Actin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 8890 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher β actin
    Liver, 22-month-old male C57BL/6J mouse. Hepatoblastoma (left) and adjacent hepatocellular adenoma (right). <t>β-catenin</t> immunohistochemistry reveals a strong nucleocytoplasmic signal in hepatoblastoma tumor cells, while neoplastic hepatocytes in the surrounding adenoma display a delicate membranous staining pattern. β-catenin immunohistochemistry (IHC); hematoxylin counterstain. Scale bar: 100 µm.
    β Actin, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 18135 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti β actin
    Liver, 22-month-old male C57BL/6J mouse. Hepatoblastoma (left) and adjacent hepatocellular adenoma (right). <t>β-catenin</t> immunohistochemistry reveals a strong nucleocytoplasmic signal in hepatoblastoma tumor cells, while neoplastic hepatocytes in the surrounding adenoma display a delicate membranous staining pattern. β-catenin immunohistochemistry (IHC); hematoxylin counterstain. Scale bar: 100 µm.
    Anti β Actin, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 8076 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore β tubulin
    Liver, 22-month-old male C57BL/6J mouse. Hepatoblastoma (left) and adjacent hepatocellular adenoma (right). <t>β-catenin</t> immunohistochemistry reveals a strong nucleocytoplasmic signal in hepatoblastoma tumor cells, while neoplastic hepatocytes in the surrounding adenoma display a delicate membranous staining pattern. β-catenin immunohistochemistry (IHC); hematoxylin counterstain. Scale bar: 100 µm.
    β Tubulin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 6958 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore igg
    Repeated Plasmodium vivax exposures increase the levels of antigen-specific <t>IgG.</t> A-C. Components of humoral response were measured in patients acutely infected for the first time (black squares) or with multiple infections (blue squares) with P . vivax . A. Pv AMA-1 and PvMSP-1 19 <t>IgM</t> and IgG were measure in plasma. B. Scattered plots showing frequency of activated memory (CD27 + CD21 - ), classical memory (CD27 + CD21 + ), naïve (CD27 - CD21 + ) and atypical memory (CD27 - CD21 - ) B cells in P . vivax -infected patients described above. C. Scattered plots showing the proportion of plasma cells (CD21 - CD20 - ) and IgG, CD38, Ki67, PD-1, IgG and PD-1 and Ki67 and CD38 expressing plasma cells from P . vivax -infected patients. All the B cell subsets were analyzed after gating on live CD19 + cells. D. Frequency of Tfh cells (PD-1 + ICOS + CXCR5 + CD45RO + CD4 + CD3 + ) cells are shown in patients infected for the first time (black squares) or infected 2 to 5 times (grey squares) or more than 5 times (blue squares) with P . vivax . Lines represent median values of the given measurement in each group (left graph). Correlation between number of malaria episodes and proportion of Tfh cells (right graph). p values are depicted in the figure.
    Igg, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 10101 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Bio-Rad β mercaptoethanol
    Repeated Plasmodium vivax exposures increase the levels of antigen-specific <t>IgG.</t> A-C. Components of humoral response were measured in patients acutely infected for the first time (black squares) or with multiple infections (blue squares) with P . vivax . A. Pv AMA-1 and PvMSP-1 19 <t>IgM</t> and IgG were measure in plasma. B. Scattered plots showing frequency of activated memory (CD27 + CD21 - ), classical memory (CD27 + CD21 + ), naïve (CD27 - CD21 + ) and atypical memory (CD27 - CD21 - ) B cells in P . vivax -infected patients described above. C. Scattered plots showing the proportion of plasma cells (CD21 - CD20 - ) and IgG, CD38, Ki67, PD-1, IgG and PD-1 and Ki67 and CD38 expressing plasma cells from P . vivax -infected patients. All the B cell subsets were analyzed after gating on live CD19 + cells. D. Frequency of Tfh cells (PD-1 + ICOS + CXCR5 + CD45RO + CD4 + CD3 + ) cells are shown in patients infected for the first time (black squares) or infected 2 to 5 times (grey squares) or more than 5 times (blue squares) with P . vivax . Lines represent median values of the given measurement in each group (left graph). Correlation between number of malaria episodes and proportion of Tfh cells (right graph). p values are depicted in the figure.
    β Mercaptoethanol, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 99/100, based on 5867 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Immunohistochemical analysis of <t>β-catenin</t> and c-Myc protein expression in the venous limb of the murine AVF 1 wk after the creation of the AVF. Immunohistochemical localization of β-catenin protein in control vein ( A ) and the venous segment
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    CCP6 or CCP1 deficiency promotes cell reprogramming. a Ccp1 - or Ccp6 - deficient MEFs were lyzed for immunoblotting. β-actin was used as a loading control. b WT or CCPs-deficient MEFs were infected by OSKM factors containing retrovirus and cultured in ESC media for 3 weeks. Alkaline phosphatase (AP)-positive colony numbers per 10 4 cells were calculated and shown as means ± S.D. **, P

    Journal: Nature Communications

    Article Title: Klf4 glutamylation is required for cell reprogramming and early embryonic development in mice

    doi: 10.1038/s41467-018-03008-2

    Figure Lengend Snippet: CCP6 or CCP1 deficiency promotes cell reprogramming. a Ccp1 - or Ccp6 - deficient MEFs were lyzed for immunoblotting. β-actin was used as a loading control. b WT or CCPs-deficient MEFs were infected by OSKM factors containing retrovirus and cultured in ESC media for 3 weeks. Alkaline phosphatase (AP)-positive colony numbers per 10 4 cells were calculated and shown as means ± S.D. **, P

    Article Snippet: Antibodies against Flag-tag (M2, F3165), β-actin (A-5316), His-tag (H1029), glutamylated tubulin (B3), and GFP-tag (G-1544) were from Sigma-Aldrich (St. Louis, USA).

    Techniques: Infection, Cell Culture

    Activation of the AKT- IκB kinase (IKK)-inhibitors of NF-κB (IκB) pathway is associated with the CK2 down-regulation-mediated nuclear import of NF-κB. ( A ) MCF-7 and HCT116 cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days. The level of each protein was determined by immunoblot analysis using specific antibodies ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of p-AKT, IKK, and IκBα relative to β-actin and that of p-IKK and p-IκBα relative to the unphosphorylated proteins ( right ). ( B ) Cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days in the absence or presence of 1 μM triciribine (TCN). Cytoplasm and nuclei were isolated from cells, and both extracts were visualized by immunoblotting. α-Tubulin (cytoplasmic marker) and histone H3 (nuclear marker) were quantified as loading controls ( left ). Representative data from three independent experiments are shown. Graphs represent the quantitation of RelA/p65 relative to the subcellular markers ( right ). exp, exposure. Data are mean ± SEM. * p

    Journal: International Journal of Molecular Sciences

    Article Title: CK2 Down-Regulation Increases the Expression of Senescence-Associated Secretory Phenotype Factors through NF-κB Activation

    doi: 10.3390/ijms22010406

    Figure Lengend Snippet: Activation of the AKT- IκB kinase (IKK)-inhibitors of NF-κB (IκB) pathway is associated with the CK2 down-regulation-mediated nuclear import of NF-κB. ( A ) MCF-7 and HCT116 cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days. The level of each protein was determined by immunoblot analysis using specific antibodies ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of p-AKT, IKK, and IκBα relative to β-actin and that of p-IKK and p-IκBα relative to the unphosphorylated proteins ( right ). ( B ) Cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days in the absence or presence of 1 μM triciribine (TCN). Cytoplasm and nuclei were isolated from cells, and both extracts were visualized by immunoblotting. α-Tubulin (cytoplasmic marker) and histone H3 (nuclear marker) were quantified as loading controls ( left ). Representative data from three independent experiments are shown. Graphs represent the quantitation of RelA/p65 relative to the subcellular markers ( right ). exp, exposure. Data are mean ± SEM. * p

    Article Snippet: Materials Antibodies against SIRT1, CK2α, p53-p21Cip1/WAF1, RelA/p65, IκB, α-tubulin, and β-actin were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

    Techniques: Activation Assay, Transfection, Quantitation Assay, Isolation, Marker

    Antisense inhibitors of miR-186, miR-216b, miR-337-3p, and miR-760 attenuate the induction of senescence-associated secretory phenotype (SASP) factors by lipopolysaccharide (LPS). ( A ) MCF-7 and HCT116 cells were transfected with the 4 miRs (miR-186, miR-216b, miR-337-3p, and miR-760) or the 4 miRi. ( B , C ) Cells were treated with LPS (6 μg/μL) in the presence or absence of the 4 miRi or pcDNA3.1-HA-CK2α for 2 days. The level of each protein was determined by immunoblot analysis using specific antibodies ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of p-AKT, IKK, and IκBα relative to β-actin and that of p-IKK and p-IκBα relative to the unphosphorylated proteins ( right ). Data are mean ± SEM. * p

    Journal: International Journal of Molecular Sciences

    Article Title: CK2 Down-Regulation Increases the Expression of Senescence-Associated Secretory Phenotype Factors through NF-κB Activation

    doi: 10.3390/ijms22010406

    Figure Lengend Snippet: Antisense inhibitors of miR-186, miR-216b, miR-337-3p, and miR-760 attenuate the induction of senescence-associated secretory phenotype (SASP) factors by lipopolysaccharide (LPS). ( A ) MCF-7 and HCT116 cells were transfected with the 4 miRs (miR-186, miR-216b, miR-337-3p, and miR-760) or the 4 miRi. ( B , C ) Cells were treated with LPS (6 μg/μL) in the presence or absence of the 4 miRi or pcDNA3.1-HA-CK2α for 2 days. The level of each protein was determined by immunoblot analysis using specific antibodies ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of p-AKT, IKK, and IκBα relative to β-actin and that of p-IKK and p-IκBα relative to the unphosphorylated proteins ( right ). Data are mean ± SEM. * p

    Article Snippet: Materials Antibodies against SIRT1, CK2α, p53-p21Cip1/WAF1, RelA/p65, IκB, α-tubulin, and β-actin were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

    Techniques: Transfection, Quantitation Assay

    CK2 down-regulation stimulates the expression of senescence-associated secretory phenotype (SASP) factors by enhancing the nuclear localization of NF-kB in human cancer cells. MCF-7 and HCT116 cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days. ( A ) The level of each mRNA was measured by RT-PCR using gene-specific primers ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of each mRNA relative to β-actin ( right ). ( B ) The level of each protein was determined by immunoblot analysis using specific antibodies ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of each protein relative to β-actin ( right ). ( C ) Cytoplasm and nuclei were isolated from cells, and both extracts were visualized by immunoblotting. α-Tubulin (cytoplasmic marker) and histone H3 (nuclear marker) were quantified as loading controls ( left ). Representative data from three independent experiments are shown. Graphs represent the quantitation of RelA/p65 relative to the subcellular markers ( right ). exp, exposure. Data are mean ± standard error of the mean (SEM). * p

    Journal: International Journal of Molecular Sciences

    Article Title: CK2 Down-Regulation Increases the Expression of Senescence-Associated Secretory Phenotype Factors through NF-κB Activation

    doi: 10.3390/ijms22010406

    Figure Lengend Snippet: CK2 down-regulation stimulates the expression of senescence-associated secretory phenotype (SASP) factors by enhancing the nuclear localization of NF-kB in human cancer cells. MCF-7 and HCT116 cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days. ( A ) The level of each mRNA was measured by RT-PCR using gene-specific primers ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of each mRNA relative to β-actin ( right ). ( B ) The level of each protein was determined by immunoblot analysis using specific antibodies ( left ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of each protein relative to β-actin ( right ). ( C ) Cytoplasm and nuclei were isolated from cells, and both extracts were visualized by immunoblotting. α-Tubulin (cytoplasmic marker) and histone H3 (nuclear marker) were quantified as loading controls ( left ). Representative data from three independent experiments are shown. Graphs represent the quantitation of RelA/p65 relative to the subcellular markers ( right ). exp, exposure. Data are mean ± standard error of the mean (SEM). * p

    Article Snippet: Materials Antibodies against SIRT1, CK2α, p53-p21Cip1/WAF1, RelA/p65, IκB, α-tubulin, and β-actin were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

    Techniques: Expressing, Transfection, Reverse Transcription Polymerase Chain Reaction, Quantitation Assay, Isolation, Marker

    SIRT1 attenuates both RelA/p65 acetylation and activation of the AKT-IKK-IκB axis mediated by CK2 down-regulation. ( A ) MCF-7 and HCT116 cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days in the absence or presence of 20 μM resveratrol or 15 mM nicotine amide. The level of each protein was determined by immunoblot analysis using specific antibodies ( top ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of acetylated p65 (ac-p65) relative to unacetylated p-65 ( bottom ). RSV, resveratrol; NA, nicotine amide. ( B ) Cells were transfected with CK2α siRNA and/or pECE-Flag-SIRT1 for two days. The level of each protein was determined by immunoblot analysis using specific antibodies ( top ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of p-AKT, IKK, and IκBα relative to β-actin and that of p-IKK and p-IκBα relative to unphosphorylated proteins ( bottom ). Data are mean ± SEM. * p

    Journal: International Journal of Molecular Sciences

    Article Title: CK2 Down-Regulation Increases the Expression of Senescence-Associated Secretory Phenotype Factors through NF-κB Activation

    doi: 10.3390/ijms22010406

    Figure Lengend Snippet: SIRT1 attenuates both RelA/p65 acetylation and activation of the AKT-IKK-IκB axis mediated by CK2 down-regulation. ( A ) MCF-7 and HCT116 cells were transfected with CK2α siRNA or pcDNA3.1-HA-CK2α for two days in the absence or presence of 20 μM resveratrol or 15 mM nicotine amide. The level of each protein was determined by immunoblot analysis using specific antibodies ( top ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of acetylated p65 (ac-p65) relative to unacetylated p-65 ( bottom ). RSV, resveratrol; NA, nicotine amide. ( B ) Cells were transfected with CK2α siRNA and/or pECE-Flag-SIRT1 for two days. The level of each protein was determined by immunoblot analysis using specific antibodies ( top ). Representative data from three independent experiments are shown. β-Actin was used as a control. Graphs represent the quantitation of p-AKT, IKK, and IκBα relative to β-actin and that of p-IKK and p-IκBα relative to unphosphorylated proteins ( bottom ). Data are mean ± SEM. * p

    Article Snippet: Materials Antibodies against SIRT1, CK2α, p53-p21Cip1/WAF1, RelA/p65, IκB, α-tubulin, and β-actin were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

    Techniques: Activation Assay, Transfection, Quantitation Assay

    Western blot representative of omentin in diabetic animals. (A) in tissues: Molecular Weight Marker (M); Mesenteric Adipose Tissue (MES), Retroperitoneal (RET), Epididimal (EPI), Brown Adipose Tissue (BAT), Liver, and Serum. For this result, 3 independent replicates were performed. (B) Representative membrane with 4 sample of omentin in muscle (one per group) compared to MES, showed that omentin was not found in muscle (I = 6 sample per group). (C) Serum Omentin Values (ELISA) ( n = 10 sample per group). (D) Omentin (40 kDa) in MES in the experimental protocols. C, control group; RT, resistance training; AT, aerobic training; CT, combined training. Results are presented as the relative density after normalizing with β-actin protein. Data are expressed as means ± SD ( n = 6 sample per group). P

    Journal: Frontiers in Physiology

    Article Title: Exercise and Omentin: Their Role in the Crosstalk Between Muscle and Adipose Tissues in Type 2 Diabetes Mellitus Rat Models

    doi: 10.3389/fphys.2018.01881

    Figure Lengend Snippet: Western blot representative of omentin in diabetic animals. (A) in tissues: Molecular Weight Marker (M); Mesenteric Adipose Tissue (MES), Retroperitoneal (RET), Epididimal (EPI), Brown Adipose Tissue (BAT), Liver, and Serum. For this result, 3 independent replicates were performed. (B) Representative membrane with 4 sample of omentin in muscle (one per group) compared to MES, showed that omentin was not found in muscle (I = 6 sample per group). (C) Serum Omentin Values (ELISA) ( n = 10 sample per group). (D) Omentin (40 kDa) in MES in the experimental protocols. C, control group; RT, resistance training; AT, aerobic training; CT, combined training. Results are presented as the relative density after normalizing with β-actin protein. Data are expressed as means ± SD ( n = 6 sample per group). P

    Article Snippet: Protein concentrations were normalized by using β-actin diluted 1:2,000 (Cell Signaling Technology, Beverly, MA, United States) in the visceral fat, or GAPDH diluted 1:10,000 (Abcam) in muscle. β-actin was detected with mouse peroxidase (HRP) – conjugated with a second antibody (Cell Signaling) diluted 1:3,000 in TBS-T, and GAPDH using antirabbit antibody (Cell Signaling), incubated and agitated for 1 h at room temperature.

    Techniques: Western Blot, Molecular Weight, Marker, Enzyme-linked Immunosorbent Assay

    Pellino2 ubiquitinates IRAK1 and suppress the inhibitory effects of IRAK1 on NLRP3. a Immunoblot analysis of myc and IRAK1 in lysates (Input) and immunoprecipitated (IP) myc samples from HEK293T cells transfected with myc-tagged Pellino2 and untagged IRAK1. b Immunoblot analysis of HA, IRAK1 and FLAG in lysates (Input) and IP IRAK1 samples from HEK293T cells transfected with FLAG-tagged Pellino2, FLAG-tagged Pellino2 RING mutant, untagged IRAK1, and HA-Ubiquitin. c, d Immunoblot analysis of Ubiquitin (short exposure (SE) or long exposure (LE)) and IRAK1 in lysates (Input) and IP IRAK1 samples or IRAK1 and ubq in TUBE-Ubq elution and cell lysates ( d ) from WT and Peli2 −/− BMDMs treated with 100 ng/ml of LPS for the indicated times. e Immunoblot analysis of NLRP3 and IRAK1 in lysates (Input) and IP IRAK1 samples from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. f ELISA of IL-1β (left panel) and TNF (right panel) in medium from primary WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for 3 h and then with 2.5 mM ATP or 5 mM Nigericin for 1 h. UT, untreated. g Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates of WT and Irak1 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h and 2.5 mM ATP for 1 h. h Immunoblot analysis of HA, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from HEK293T cells transfected with V5-tagged NLRP3, untagged IRAK1, untagged kinase dead IRAK1 (IRAK1-KD), and HA-ubiquitin. β-Actin was used as loading controls. i Immunoblot analysis of Ubiquitin, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from immortalized WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. j Immortalized WT and Irak1 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING) or Pellino2 FHA mutant (FHA). Immunoblot analysis of myc and NLRP3 in lysates (Input) and IP myc samples from virus-infected BMDMs treated with 100 ng/ml LPS for indicated times. * p

    Journal: Nature Communications

    Article Title: The E3 ubiquitin ligase Pellino2 mediates priming of the NLRP3 inflammasome

    doi: 10.1038/s41467-018-03669-z

    Figure Lengend Snippet: Pellino2 ubiquitinates IRAK1 and suppress the inhibitory effects of IRAK1 on NLRP3. a Immunoblot analysis of myc and IRAK1 in lysates (Input) and immunoprecipitated (IP) myc samples from HEK293T cells transfected with myc-tagged Pellino2 and untagged IRAK1. b Immunoblot analysis of HA, IRAK1 and FLAG in lysates (Input) and IP IRAK1 samples from HEK293T cells transfected with FLAG-tagged Pellino2, FLAG-tagged Pellino2 RING mutant, untagged IRAK1, and HA-Ubiquitin. c, d Immunoblot analysis of Ubiquitin (short exposure (SE) or long exposure (LE)) and IRAK1 in lysates (Input) and IP IRAK1 samples or IRAK1 and ubq in TUBE-Ubq elution and cell lysates ( d ) from WT and Peli2 −/− BMDMs treated with 100 ng/ml of LPS for the indicated times. e Immunoblot analysis of NLRP3 and IRAK1 in lysates (Input) and IP IRAK1 samples from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. f ELISA of IL-1β (left panel) and TNF (right panel) in medium from primary WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for 3 h and then with 2.5 mM ATP or 5 mM Nigericin for 1 h. UT, untreated. g Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates of WT and Irak1 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h and 2.5 mM ATP for 1 h. h Immunoblot analysis of HA, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from HEK293T cells transfected with V5-tagged NLRP3, untagged IRAK1, untagged kinase dead IRAK1 (IRAK1-KD), and HA-ubiquitin. β-Actin was used as loading controls. i Immunoblot analysis of Ubiquitin, NLRP3, and IRAK1 in lysates (Input) and IP NLRP3 samples from immortalized WT and Irak1 −/− BMDMs treated with 100 ng/ml LPS for the indicated times. j Immortalized WT and Irak1 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING) or Pellino2 FHA mutant (FHA). Immunoblot analysis of myc and NLRP3 in lysates (Input) and IP myc samples from virus-infected BMDMs treated with 100 ng/ml LPS for indicated times. * p

    Article Snippet: Anti-p-ERK (9101), anti-ERK (9102), anti-p-IκBα (9246), anti-pIKK (14938), anti-p65 (3033) anti-p-P38 (9211), anti-P38 (9212), anti-p-Jnk (9251), anti-Jnk (9252), anti-myc (2276), anti-IRAK1 (4504), anti-IRAK4 (4363), and anti-human caspase 1 p20 (4199) were from CellSignaling; anti-mouse caspase1 p20 (AG-20B-0042-C100) and anti-mouse NLRP3 (AG-20B-0014-C100) was from adipogen; anti-mouse IL1β (AF-401-NA) was from R & D; anti-IκB-α (C-21; sc-371), anti-Ub (P4D1; sc-8017), and anti-ASC (sc-514414) was from Santa Cruz; anti-HA (16B12; MMS-101P) was from Covance; anti-K63 ubiquitin (HWA4C4; BML-PW0600) was from Enzo life science; anti-K48 ubiquitin (Apu2; 05-1307) was from Millipore; anti-flag (F3165) and anti-β-actin (AC-15; A 1978) were from Sigma; anti-mouse IRDyeTM 680 (926-68070) and anti-rabbit IRDyeTM 800 (926-32211) were from LI-COR Biosciences; anti-mouse-HRP and anti-rabbit-HRP were from Promega.

    Techniques: Immunoprecipitation, Transfection, Mutagenesis, Enzyme-linked Immunosorbent Assay, Infection, Expressing, Construct

    Pellino2 mediates NLRP3-dependent oligomerization of ASC. a Immunofluorescence staining of ASC in WT and Peli2 −/− BMDMs that were left untreated (UT) or treated with 100 ng/ml LPS for 3 h and further stimulated with ATP for 30 min. ASC specks were detected by immunostaining using anti-ASC antibody and anti-rabbit Alexa Fluor 568 (red) and cells were counter stained with nuclei-staining DAPI. The histogram quantitates the percentage of cells that exhibit ASC speck formation. (scale bar = 100 μm). b, c Immunoblot analysis of ASC in chemically cross-linked NP-40 insoluble fractions and in NP-40 soluble fractions from cell lysates of WT and Peli2 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h with or without further treatment with b 2.5 mM ATP, or c 5 mM Nigericin for 30 min. β-Actin was used as loading controls. d WT and Peli2 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING), or Pellino2 FHA mutant (FHA). Immunoblot analysis of ASC in chemically cross-linked NP-40 insoluble fractions and in NP-40 soluble fractions from cell lysates of MSCV-infected cells treated with 100 ng/ml LPS for 3 h followed by 2.5 mM ATP for 30 min. The expression of the Pellino2 constructs was measured by immunoblotting with an anti-myc antibody. ** p

    Journal: Nature Communications

    Article Title: The E3 ubiquitin ligase Pellino2 mediates priming of the NLRP3 inflammasome

    doi: 10.1038/s41467-018-03669-z

    Figure Lengend Snippet: Pellino2 mediates NLRP3-dependent oligomerization of ASC. a Immunofluorescence staining of ASC in WT and Peli2 −/− BMDMs that were left untreated (UT) or treated with 100 ng/ml LPS for 3 h and further stimulated with ATP for 30 min. ASC specks were detected by immunostaining using anti-ASC antibody and anti-rabbit Alexa Fluor 568 (red) and cells were counter stained with nuclei-staining DAPI. The histogram quantitates the percentage of cells that exhibit ASC speck formation. (scale bar = 100 μm). b, c Immunoblot analysis of ASC in chemically cross-linked NP-40 insoluble fractions and in NP-40 soluble fractions from cell lysates of WT and Peli2 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h with or without further treatment with b 2.5 mM ATP, or c 5 mM Nigericin for 30 min. β-Actin was used as loading controls. d WT and Peli2 −/− BMDMs were infected with MSCV as control (Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (Peli2), Pellino2 RING mutant (RING), or Pellino2 FHA mutant (FHA). Immunoblot analysis of ASC in chemically cross-linked NP-40 insoluble fractions and in NP-40 soluble fractions from cell lysates of MSCV-infected cells treated with 100 ng/ml LPS for 3 h followed by 2.5 mM ATP for 30 min. The expression of the Pellino2 constructs was measured by immunoblotting with an anti-myc antibody. ** p

    Article Snippet: Anti-p-ERK (9101), anti-ERK (9102), anti-p-IκBα (9246), anti-pIKK (14938), anti-p65 (3033) anti-p-P38 (9211), anti-P38 (9212), anti-p-Jnk (9251), anti-Jnk (9252), anti-myc (2276), anti-IRAK1 (4504), anti-IRAK4 (4363), and anti-human caspase 1 p20 (4199) were from CellSignaling; anti-mouse caspase1 p20 (AG-20B-0042-C100) and anti-mouse NLRP3 (AG-20B-0014-C100) was from adipogen; anti-mouse IL1β (AF-401-NA) was from R & D; anti-IκB-α (C-21; sc-371), anti-Ub (P4D1; sc-8017), and anti-ASC (sc-514414) was from Santa Cruz; anti-HA (16B12; MMS-101P) was from Covance; anti-K63 ubiquitin (HWA4C4; BML-PW0600) was from Enzo life science; anti-K48 ubiquitin (Apu2; 05-1307) was from Millipore; anti-flag (F3165) and anti-β-actin (AC-15; A 1978) were from Sigma; anti-mouse IRDyeTM 680 (926-68070) and anti-rabbit IRDyeTM 800 (926-32211) were from LI-COR Biosciences; anti-mouse-HRP and anti-rabbit-HRP were from Promega.

    Techniques: Immunofluorescence, Staining, Immunostaining, Infection, Expressing, Construct, Mutagenesis

    Pellino2 mediates LPS-induced ubiquitination and activation of NLRP3. a-c Immunoblot analysis of NLRP3 and ubiquitin in cell lysates (Input) and immunoprecipitated (IP) NLRP3 samples ( a ) or NLRP3 and K63-linked ubiquitin (K63-ubq) in K63-TUBE-FLAG elution and cell lysates ( b ) or NLRP3 and K48-ubq in K48-TUBE-FLAG elution and cell lysates ( c ) from WT and Peli2 −/− BMDMs treated with a 100 ng/ml LPS for the indicated times. d ELISA of IL-1β in medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 2 h followed by sequential treatment with 1 μM MCC950 for 1 h and 2.5 mM ATP for 30 min. UT, untreated. e, f Immunoblot analysis of ubiquitin and NLRP3 in lysates (Input) and immunoprecipitated (IP) NLRP3 samples ( e ) or NLRP3 and K63-ubq in K63-TUBE-FLAG elution and cell lysates ( f ) from WT and Peli2 −/− BMDMs pre-treated with 1 μM MCC950 for 1 h followed by treatment with 100 ng/ml LPS for the indicated times. g Peli2 −/− BMDMs were infected with MSCV as control (MSCV-Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (MSCV-Peli2). Immunoblot analysis of NLRP3 and myc in lysates (Input) and immunoprecipitated (IP) myc samples from MSCV-infected cells treated with 100 ng/ml LPS for the indicated times. β-Actin was used as loading controls. * p

    Journal: Nature Communications

    Article Title: The E3 ubiquitin ligase Pellino2 mediates priming of the NLRP3 inflammasome

    doi: 10.1038/s41467-018-03669-z

    Figure Lengend Snippet: Pellino2 mediates LPS-induced ubiquitination and activation of NLRP3. a-c Immunoblot analysis of NLRP3 and ubiquitin in cell lysates (Input) and immunoprecipitated (IP) NLRP3 samples ( a ) or NLRP3 and K63-linked ubiquitin (K63-ubq) in K63-TUBE-FLAG elution and cell lysates ( b ) or NLRP3 and K48-ubq in K48-TUBE-FLAG elution and cell lysates ( c ) from WT and Peli2 −/− BMDMs treated with a 100 ng/ml LPS for the indicated times. d ELISA of IL-1β in medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 2 h followed by sequential treatment with 1 μM MCC950 for 1 h and 2.5 mM ATP for 30 min. UT, untreated. e, f Immunoblot analysis of ubiquitin and NLRP3 in lysates (Input) and immunoprecipitated (IP) NLRP3 samples ( e ) or NLRP3 and K63-ubq in K63-TUBE-FLAG elution and cell lysates ( f ) from WT and Peli2 −/− BMDMs pre-treated with 1 μM MCC950 for 1 h followed by treatment with 100 ng/ml LPS for the indicated times. g Peli2 −/− BMDMs were infected with MSCV as control (MSCV-Ctrl) or with MSCV containing an expression construct encoding myc-tagged murine Pellino2 (MSCV-Peli2). Immunoblot analysis of NLRP3 and myc in lysates (Input) and immunoprecipitated (IP) myc samples from MSCV-infected cells treated with 100 ng/ml LPS for the indicated times. β-Actin was used as loading controls. * p

    Article Snippet: Anti-p-ERK (9101), anti-ERK (9102), anti-p-IκBα (9246), anti-pIKK (14938), anti-p65 (3033) anti-p-P38 (9211), anti-P38 (9212), anti-p-Jnk (9251), anti-Jnk (9252), anti-myc (2276), anti-IRAK1 (4504), anti-IRAK4 (4363), and anti-human caspase 1 p20 (4199) were from CellSignaling; anti-mouse caspase1 p20 (AG-20B-0042-C100) and anti-mouse NLRP3 (AG-20B-0014-C100) was from adipogen; anti-mouse IL1β (AF-401-NA) was from R & D; anti-IκB-α (C-21; sc-371), anti-Ub (P4D1; sc-8017), and anti-ASC (sc-514414) was from Santa Cruz; anti-HA (16B12; MMS-101P) was from Covance; anti-K63 ubiquitin (HWA4C4; BML-PW0600) was from Enzo life science; anti-K48 ubiquitin (Apu2; 05-1307) was from Millipore; anti-flag (F3165) and anti-β-actin (AC-15; A 1978) were from Sigma; anti-mouse IRDyeTM 680 (926-68070) and anti-rabbit IRDyeTM 800 (926-32211) were from LI-COR Biosciences; anti-mouse-HRP and anti-rabbit-HRP were from Promega.

    Techniques: Activation Assay, Immunoprecipitation, Enzyme-linked Immunosorbent Assay, Infection, Expressing, Construct

    Pellino2 mediates activation of the NLRP3 pathway. ELISA of a IL-1β and b IL-18, and c LDH assay of medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 3 h with or without further stimulation with 2.5 mM ATP for 1 h. UT, untreated. d Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates from WT and Peli2 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h with or without further stimulation with 2.5 mM ATP for 1 h or transfection of Poly (dA:dT) (1 μg/ml) for 6 h. ELISA of e IL-1β and f IL-18 and g LDH assay of medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 3 h with or without further stimulation with 5 mM Nigericin for 1 h. h Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates from WT and Peli2 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h with or without further stimulation with 5 mM Nigericin for 1 h or transfection of Poly (dA:dT) (1 μg/ml) for 6 h. i ELISA of IL-1β of medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 3 h with or without further stimulation with the indicated concentrations of Alum for 6 h. j ELISA of IL-1β in medium from peritoneal-resident macrophages isolated from WT and Peli2 −/− mice. Cells were treated with 100 ng/ml of LPS for 3 h with or without further stimulation with 2.5 mM ATP for 1 h. k-m Human THP1 cells were transfected with human Pellino2-specific siRNA or control siRNA. k Quantitative RT-PCR analysis of PELI2 expression in transfected cells. l ELISA of IL-1β in medium from transfected THP1 cells stimulated with 100 ng/ml LPS for 6 h with or without further treatment with 2.5 mM ATP or 5 mM Nigericin for 1 h. m Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates from transfected THP1 cells stimulated with 100 ng/ml LPS for 6 h with or without further stimulation with 2.5 mM ATP or 5 mM Nigericin for 1 h. β-Actin was used as loading controls. * p

    Journal: Nature Communications

    Article Title: The E3 ubiquitin ligase Pellino2 mediates priming of the NLRP3 inflammasome

    doi: 10.1038/s41467-018-03669-z

    Figure Lengend Snippet: Pellino2 mediates activation of the NLRP3 pathway. ELISA of a IL-1β and b IL-18, and c LDH assay of medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 3 h with or without further stimulation with 2.5 mM ATP for 1 h. UT, untreated. d Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates from WT and Peli2 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h with or without further stimulation with 2.5 mM ATP for 1 h or transfection of Poly (dA:dT) (1 μg/ml) for 6 h. ELISA of e IL-1β and f IL-18 and g LDH assay of medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 3 h with or without further stimulation with 5 mM Nigericin for 1 h. h Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates from WT and Peli2 −/− BMDMs stimulated with 100 ng/ml LPS for 3 h with or without further stimulation with 5 mM Nigericin for 1 h or transfection of Poly (dA:dT) (1 μg/ml) for 6 h. i ELISA of IL-1β of medium from WT and Peli2 −/− BMDMs treated with 100 ng/ml LPS for 3 h with or without further stimulation with the indicated concentrations of Alum for 6 h. j ELISA of IL-1β in medium from peritoneal-resident macrophages isolated from WT and Peli2 −/− mice. Cells were treated with 100 ng/ml of LPS for 3 h with or without further stimulation with 2.5 mM ATP for 1 h. k-m Human THP1 cells were transfected with human Pellino2-specific siRNA or control siRNA. k Quantitative RT-PCR analysis of PELI2 expression in transfected cells. l ELISA of IL-1β in medium from transfected THP1 cells stimulated with 100 ng/ml LPS for 6 h with or without further treatment with 2.5 mM ATP or 5 mM Nigericin for 1 h. m Immunoblot analysis of IL-1β and Caspase-1 in medium (Sup) and lysates from transfected THP1 cells stimulated with 100 ng/ml LPS for 6 h with or without further stimulation with 2.5 mM ATP or 5 mM Nigericin for 1 h. β-Actin was used as loading controls. * p

    Article Snippet: Anti-p-ERK (9101), anti-ERK (9102), anti-p-IκBα (9246), anti-pIKK (14938), anti-p65 (3033) anti-p-P38 (9211), anti-P38 (9212), anti-p-Jnk (9251), anti-Jnk (9252), anti-myc (2276), anti-IRAK1 (4504), anti-IRAK4 (4363), and anti-human caspase 1 p20 (4199) were from CellSignaling; anti-mouse caspase1 p20 (AG-20B-0042-C100) and anti-mouse NLRP3 (AG-20B-0014-C100) was from adipogen; anti-mouse IL1β (AF-401-NA) was from R & D; anti-IκB-α (C-21; sc-371), anti-Ub (P4D1; sc-8017), and anti-ASC (sc-514414) was from Santa Cruz; anti-HA (16B12; MMS-101P) was from Covance; anti-K63 ubiquitin (HWA4C4; BML-PW0600) was from Enzo life science; anti-K48 ubiquitin (Apu2; 05-1307) was from Millipore; anti-flag (F3165) and anti-β-actin (AC-15; A 1978) were from Sigma; anti-mouse IRDyeTM 680 (926-68070) and anti-rabbit IRDyeTM 800 (926-32211) were from LI-COR Biosciences; anti-mouse-HRP and anti-rabbit-HRP were from Promega.

    Techniques: Activation Assay, Enzyme-linked Immunosorbent Assay, Lactate Dehydrogenase Assay, Transfection, Isolation, Mouse Assay, Quantitative RT-PCR, Expressing

    Pellino2 mediates activation of the NLRP3 pathway in response to bacterial infection. a-f WT and Peli2 −/− BMDMs were infected with a, b C. rodentium , c, d E.coli , or e, f P. aeruginosa (PA01 strain) at a multiplicity of infection (MOI) of 100. a, c, e ELISA of IL-1β, IL-18, and CXCL1 in medium from BMDMs infected for 6 h. b, d Immunoblot analysis of Caspase-11 in lysates from cells infected for 0–6 h. f Immunoblot analysis of IL-1β and Caspase-1 in lysates from cells infected with PAO1 for 3 h. β-Actin was used as loading controls. g ELISA of IL-1β, IL-18, and IL-6 in peritoneal lavage from WT and Peli2 −/− mice previously infected for 10 h by intraperitoneal injection of PAO1 (1.5 × 10 7 CFU). * p

    Journal: Nature Communications

    Article Title: The E3 ubiquitin ligase Pellino2 mediates priming of the NLRP3 inflammasome

    doi: 10.1038/s41467-018-03669-z

    Figure Lengend Snippet: Pellino2 mediates activation of the NLRP3 pathway in response to bacterial infection. a-f WT and Peli2 −/− BMDMs were infected with a, b C. rodentium , c, d E.coli , or e, f P. aeruginosa (PA01 strain) at a multiplicity of infection (MOI) of 100. a, c, e ELISA of IL-1β, IL-18, and CXCL1 in medium from BMDMs infected for 6 h. b, d Immunoblot analysis of Caspase-11 in lysates from cells infected for 0–6 h. f Immunoblot analysis of IL-1β and Caspase-1 in lysates from cells infected with PAO1 for 3 h. β-Actin was used as loading controls. g ELISA of IL-1β, IL-18, and IL-6 in peritoneal lavage from WT and Peli2 −/− mice previously infected for 10 h by intraperitoneal injection of PAO1 (1.5 × 10 7 CFU). * p

    Article Snippet: Anti-p-ERK (9101), anti-ERK (9102), anti-p-IκBα (9246), anti-pIKK (14938), anti-p65 (3033) anti-p-P38 (9211), anti-P38 (9212), anti-p-Jnk (9251), anti-Jnk (9252), anti-myc (2276), anti-IRAK1 (4504), anti-IRAK4 (4363), and anti-human caspase 1 p20 (4199) were from CellSignaling; anti-mouse caspase1 p20 (AG-20B-0042-C100) and anti-mouse NLRP3 (AG-20B-0014-C100) was from adipogen; anti-mouse IL1β (AF-401-NA) was from R & D; anti-IκB-α (C-21; sc-371), anti-Ub (P4D1; sc-8017), and anti-ASC (sc-514414) was from Santa Cruz; anti-HA (16B12; MMS-101P) was from Covance; anti-K63 ubiquitin (HWA4C4; BML-PW0600) was from Enzo life science; anti-K48 ubiquitin (Apu2; 05-1307) was from Millipore; anti-flag (F3165) and anti-β-actin (AC-15; A 1978) were from Sigma; anti-mouse IRDyeTM 680 (926-68070) and anti-rabbit IRDyeTM 800 (926-32211) were from LI-COR Biosciences; anti-mouse-HRP and anti-rabbit-HRP were from Promega.

    Techniques: Activation Assay, Infection, Enzyme-linked Immunosorbent Assay, Mouse Assay, Injection

    Rescue of KIF3A expression in Kif3a -/- MEFs does not attenuate RHOA activation mediated by purmorphamine. A . GLI1 induction and KIF3A expression in WT MEFs (WT) vs. Kif3a -/- MEFs infected with AdV-Kif3a at MOIs of 10 and 80. p38 was used as loading control (n = 3). B . Staining of primary cilia in WT MEFs, Kif3a -/- MEFs, or Kif3a -/- MEFs infected with either control (empty) AdV or AdV-Kif3a (MOI = 10). Cells were stained with anti-acetylated α-tubulin and anti-Alexa 488 to visualize the axoneme of each primary cilium. Quantification of the percentage of cilia-positive cells (mean ± S.E.M; n = 4) C . RHOA activation in Kif3a -/- MEFs infected with control AdV or AdV-Kif3a at MOI = 10 and stimulated with 5 μM PUR for the indicated times (β-actin was used as loading control because total RHOA was obscured by a cross-reactive protein that appears as a consequence of AdV infection). D . Densitometric quantification of RHOA-GTP/ β-actin increase in response to 5 μM PUR at the indicated times. The ratios were expressed as fold change compared to t = 0. * p

    Journal: PLoS ONE

    Article Title: Activation of the Gi protein-RHOA axis by non-canonical Hedgehog signaling is independent of primary cilia

    doi: 10.1371/journal.pone.0203170

    Figure Lengend Snippet: Rescue of KIF3A expression in Kif3a -/- MEFs does not attenuate RHOA activation mediated by purmorphamine. A . GLI1 induction and KIF3A expression in WT MEFs (WT) vs. Kif3a -/- MEFs infected with AdV-Kif3a at MOIs of 10 and 80. p38 was used as loading control (n = 3). B . Staining of primary cilia in WT MEFs, Kif3a -/- MEFs, or Kif3a -/- MEFs infected with either control (empty) AdV or AdV-Kif3a (MOI = 10). Cells were stained with anti-acetylated α-tubulin and anti-Alexa 488 to visualize the axoneme of each primary cilium. Quantification of the percentage of cilia-positive cells (mean ± S.E.M; n = 4) C . RHOA activation in Kif3a -/- MEFs infected with control AdV or AdV-Kif3a at MOI = 10 and stimulated with 5 μM PUR for the indicated times (β-actin was used as loading control because total RHOA was obscured by a cross-reactive protein that appears as a consequence of AdV infection). D . Densitometric quantification of RHOA-GTP/ β-actin increase in response to 5 μM PUR at the indicated times. The ratios were expressed as fold change compared to t = 0. * p

    Article Snippet: The β-actin antibody (Cat#A1978 clone AC-15) was purchased from Sigma-Aldrich and used at 1:10000–1:40000 dilution.

    Techniques: Expressing, Activation Assay, Infection, Staining

    Staufen1 knockdown inhibits Prostate Cancer cell migration. Migration assays were performed after 48 h of Staufen1 knockdown and a is a representative western blot of Staufen1 expression with β-actin as a loading control in PC3 and DU145 cells. Each representative blot is cropped to show an n = 1 for each cell line from their respective full-length blot. Note that the DU145 cells used for Fig. 4 were also used for Fig. 5 experiments and therefore a different representative image from the same blot was selected for each figure. Quantification is represented normalized to CTL (n = 3). Data are Mean ± SD, One-Sample T-Test, **P

    Journal: BMC Cancer

    Article Title: Distinct roles for the RNA-binding protein Staufen1 in prostate cancer

    doi: 10.1186/s12885-021-07844-2

    Figure Lengend Snippet: Staufen1 knockdown inhibits Prostate Cancer cell migration. Migration assays were performed after 48 h of Staufen1 knockdown and a is a representative western blot of Staufen1 expression with β-actin as a loading control in PC3 and DU145 cells. Each representative blot is cropped to show an n = 1 for each cell line from their respective full-length blot. Note that the DU145 cells used for Fig. 4 were also used for Fig. 5 experiments and therefore a different representative image from the same blot was selected for each figure. Quantification is represented normalized to CTL (n = 3). Data are Mean ± SD, One-Sample T-Test, **P

    Article Snippet: The antibodies used were anti-Staufen1 (ab73478, Abcam, Ontario, Canada), anti-phospho (Ser2448)-mTOR (#2971, Cell Signaling Technology, Danvers, MA, USA), anti-mTOR (#2983, Cell Signaling Technology, Danvers, MA, USA), anti-phospho (Try576/577)-FAK (#3281, Cell Signaling Technology, Danvers, MA, USA), anti-phospho (Tyr397)-FAK (#8556, Cell Signaling Technology, Danvers, MA, USA), anti-FAK (#13009, Cell Signaling Technology, Danvers, MA, USA), anti-phospho-4E-BP1 (Thr37/46) (# 9459, Cell Signaling Technology, Danvers, MA, USA), anti-4E-BP1 (# 9452, Cell Signaling Technology, Danvers, MA, USA), HA.11 clone 16B12 (1:1000; BioLegend, California, USA), anti-β-actin (#47778, Santa Cruz Biotechnology, CA, USA), and anti-GAPDH (ab8245, Abcam, Ontario, Canada).

    Techniques: Migration, Western Blot, Expressing

    Staufen1 Differentially Regulates Tumorigenesis via regulation of Signaling Pathways across Prostate Cancer Cell Lines. Western blot analysis was performed 48 h post-infection of prostate cancer cells with Control (CTL) or Staufen1-shRNA (shStau1) lentivirus ( a and d ) representative Staufen1 expression in LNCaP and PC3 cells, respectively. LNCaP cells were examined for ( b ) expression of total mTOR and phospho-mTOR (Ser2448) and ( c ) expression of total 4E-BP1 and phospho-4E-BP1 (Thr37/46). PC3 cells were analyzed for members of the FAK signaling pathway where ( e ) represents total FAK and phospho-FAK (Tyr397 and Tyr576/577). All quantifications are normalized to loading controls GAPDH or β-actin and represented as a fold change relative to the Control (CTL) with n = 4. Data are Mean ± SD, One-Sample T-Test, *P

    Journal: BMC Cancer

    Article Title: Distinct roles for the RNA-binding protein Staufen1 in prostate cancer

    doi: 10.1186/s12885-021-07844-2

    Figure Lengend Snippet: Staufen1 Differentially Regulates Tumorigenesis via regulation of Signaling Pathways across Prostate Cancer Cell Lines. Western blot analysis was performed 48 h post-infection of prostate cancer cells with Control (CTL) or Staufen1-shRNA (shStau1) lentivirus ( a and d ) representative Staufen1 expression in LNCaP and PC3 cells, respectively. LNCaP cells were examined for ( b ) expression of total mTOR and phospho-mTOR (Ser2448) and ( c ) expression of total 4E-BP1 and phospho-4E-BP1 (Thr37/46). PC3 cells were analyzed for members of the FAK signaling pathway where ( e ) represents total FAK and phospho-FAK (Tyr397 and Tyr576/577). All quantifications are normalized to loading controls GAPDH or β-actin and represented as a fold change relative to the Control (CTL) with n = 4. Data are Mean ± SD, One-Sample T-Test, *P

    Article Snippet: The antibodies used were anti-Staufen1 (ab73478, Abcam, Ontario, Canada), anti-phospho (Ser2448)-mTOR (#2971, Cell Signaling Technology, Danvers, MA, USA), anti-mTOR (#2983, Cell Signaling Technology, Danvers, MA, USA), anti-phospho (Try576/577)-FAK (#3281, Cell Signaling Technology, Danvers, MA, USA), anti-phospho (Tyr397)-FAK (#8556, Cell Signaling Technology, Danvers, MA, USA), anti-FAK (#13009, Cell Signaling Technology, Danvers, MA, USA), anti-phospho-4E-BP1 (Thr37/46) (# 9459, Cell Signaling Technology, Danvers, MA, USA), anti-4E-BP1 (# 9452, Cell Signaling Technology, Danvers, MA, USA), HA.11 clone 16B12 (1:1000; BioLegend, California, USA), anti-β-actin (#47778, Santa Cruz Biotechnology, CA, USA), and anti-GAPDH (ab8245, Abcam, Ontario, Canada).

    Techniques: Western Blot, Infection, shRNA, Expressing

    Staufen1 knockdown inhibits Prostate Cancer cell invasion and motility. Motility and Invasion assays were performed following 48 h of Control (CTL) or Staufen1-shRNA (shStau1) expression. a Western blot analysis of Staufen1 expression showing a representative blot of Staufen1 with β-actin as a loading control in PC3 and DU145 cells. Each representative blot is cropped to show an n = 1 for each cell line from their respective full-length blot. Note that the DU145 cells used for Fig. 4 were also used for Fig. 5 experiments and therefore a different representative image from the same blot was selected for each figure. Quantification of n = 4 is represented normalized to CTL. Data are Mean ± SD, One-Sample T-Test, **P

    Journal: BMC Cancer

    Article Title: Distinct roles for the RNA-binding protein Staufen1 in prostate cancer

    doi: 10.1186/s12885-021-07844-2

    Figure Lengend Snippet: Staufen1 knockdown inhibits Prostate Cancer cell invasion and motility. Motility and Invasion assays were performed following 48 h of Control (CTL) or Staufen1-shRNA (shStau1) expression. a Western blot analysis of Staufen1 expression showing a representative blot of Staufen1 with β-actin as a loading control in PC3 and DU145 cells. Each representative blot is cropped to show an n = 1 for each cell line from their respective full-length blot. Note that the DU145 cells used for Fig. 4 were also used for Fig. 5 experiments and therefore a different representative image from the same blot was selected for each figure. Quantification of n = 4 is represented normalized to CTL. Data are Mean ± SD, One-Sample T-Test, **P

    Article Snippet: The antibodies used were anti-Staufen1 (ab73478, Abcam, Ontario, Canada), anti-phospho (Ser2448)-mTOR (#2971, Cell Signaling Technology, Danvers, MA, USA), anti-mTOR (#2983, Cell Signaling Technology, Danvers, MA, USA), anti-phospho (Try576/577)-FAK (#3281, Cell Signaling Technology, Danvers, MA, USA), anti-phospho (Tyr397)-FAK (#8556, Cell Signaling Technology, Danvers, MA, USA), anti-FAK (#13009, Cell Signaling Technology, Danvers, MA, USA), anti-phospho-4E-BP1 (Thr37/46) (# 9459, Cell Signaling Technology, Danvers, MA, USA), anti-4E-BP1 (# 9452, Cell Signaling Technology, Danvers, MA, USA), HA.11 clone 16B12 (1:1000; BioLegend, California, USA), anti-β-actin (#47778, Santa Cruz Biotechnology, CA, USA), and anti-GAPDH (ab8245, Abcam, Ontario, Canada).

    Techniques: shRNA, Expressing, Western Blot

    Effect of CMDA on the phosphorylation of MAPK and AP-1. (A) Effect of CMDA on the total and phosphorylated protein levels of p38, ERK and JNK MAPKs was analyzed in UVA-irradiated (10 J/cm 2 ) HaCaT keratinocytes after 24 h of incubation via western blotting. β-actin was used as an internal loading control. (B) Effect of CMDA on the total and phosphorylated protein levels of c-Fos and c-Jun was investigated in both total cell lysates and nuclear fractions of UVA-irradiated (10 J/cm 2 ) HaCaT keratinocytes after 24 h of incubation via western blotting. β-actin and lamin B1 were used as internal loading controls. CMDA, camellioside A; MAPK, mitogen-activated protein kinase; AP-1, activator protein 1; UVA, ultraviolet A.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Camellioside A, isolated from Camellia japonica flowers, attenuates UVA-induced production of MMP-1 in HaCaT keratinocytes via suppression of MAPK activation

    doi: 10.3892/etm.2020.9448

    Figure Lengend Snippet: Effect of CMDA on the phosphorylation of MAPK and AP-1. (A) Effect of CMDA on the total and phosphorylated protein levels of p38, ERK and JNK MAPKs was analyzed in UVA-irradiated (10 J/cm 2 ) HaCaT keratinocytes after 24 h of incubation via western blotting. β-actin was used as an internal loading control. (B) Effect of CMDA on the total and phosphorylated protein levels of c-Fos and c-Jun was investigated in both total cell lysates and nuclear fractions of UVA-irradiated (10 J/cm 2 ) HaCaT keratinocytes after 24 h of incubation via western blotting. β-actin and lamin B1 were used as internal loading controls. CMDA, camellioside A; MAPK, mitogen-activated protein kinase; AP-1, activator protein 1; UVA, ultraviolet A.

    Article Snippet: The following primary antibodies were used: MMP-1 (cat. no. sc-6837; Santa Cruz Biotechnology, Inc.), MMP-9 (cat. no. 393857; Cell Signaling Technology, Inc.), type I pro-collagen (cat. no. sc-8782; Santa Cruz Biotechnology, Inc.), p38 (cat. no. 8690; Cell Signaling Technology, Inc.), phosphorylated (p)-p38 (cat. no. 4511; Cell Signaling Technology, Inc.), JNK (cat. no. LF-PA0047; Thermo Fisher Scientific, Inc.), p-JNK (cat. no. sc-293136; Santa Cruz Biotechnology, Inc.), ERK (cat. no. 4695; Cell Signaling Technology, Inc.), p-ERK (cat. no. 4370; Cell Signaling Technology, Inc.), c-Jun (cat. no. sc-74543; Santa Cruz Biotechnology, Inc.), p-c-Jun (cat. no. sc-822; Santa Cruz Biotechnology, Inc.), c-Fos (cat. no. sc-7202; Santa Cruz Biotechnology, Inc.), p-c-Fos (cat. no. 5348s; Cell Signaling Technology, Inc.), β-actin (cat. no. sc-47778; Santa Cruz Biotechnology, Inc.) and lamin B1 (cat. no. sc-374015; Santa Cruz Biotechnology, Inc.).

    Techniques: Irradiation, Incubation, Western Blot

    Gly inhibited d -galactose-induced synaptic and memory dysfunction in C57BL/6 mice brain. a Western blot analysis of presynaptic proteins including synaptophysin (SYP), syntaxin (SYN), and postsynaptic density proteins (PSD95) in the hippocampus of different experimental groups The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. b The immunofluorescence images represented the immunoreactivity of PSD95 (green, FITC; blue, DAPI) in cortex and hippocampus of mice brain; along with their relative histograms, respectively. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments performed N = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions = 100 μm. c Mean escape latency in seconds to reach the hidden platform during training (5 days) along d with representative trajectories; f number of target crossings; g the time spent in the target quadrant; h Y-maze analysis represented spontaneous alteration behaviors along with its representative trajectories of mice. For behavioral study, the number of mice ( n = 16) per experimental group was used. Asterisk (*) sign indicated significant difference from the normal saline treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated group

    Journal: Journal of Neuroinflammation

    Article Title: Glycine, the smallest amino acid, confers neuroprotection against d-galactose-induced neurodegeneration and memory impairment by regulating c-Jun N-terminal kinase in the mouse brain

    doi: 10.1186/s12974-020-01989-w

    Figure Lengend Snippet: Gly inhibited d -galactose-induced synaptic and memory dysfunction in C57BL/6 mice brain. a Western blot analysis of presynaptic proteins including synaptophysin (SYP), syntaxin (SYN), and postsynaptic density proteins (PSD95) in the hippocampus of different experimental groups The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. b The immunofluorescence images represented the immunoreactivity of PSD95 (green, FITC; blue, DAPI) in cortex and hippocampus of mice brain; along with their relative histograms, respectively. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments performed N = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions = 100 μm. c Mean escape latency in seconds to reach the hidden platform during training (5 days) along d with representative trajectories; f number of target crossings; g the time spent in the target quadrant; h Y-maze analysis represented spontaneous alteration behaviors along with its representative trajectories of mice. For behavioral study, the number of mice ( n = 16) per experimental group was used. Asterisk (*) sign indicated significant difference from the normal saline treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated group

    Article Snippet: To confirm equal sample loading, an anti-β-actin antibody (Santa Cruz Biotechnology, Dallas, TX, USA) was used as a standard for comparison.

    Techniques: Mouse Assay, Western Blot, Software, Immunofluorescence, Staining

    Glycine treatment reduced d -galactose-mediated elevated p-JNK-dependent neuroapoptosis in HT22 cells lines. a Representative western blot analysis of activated phosphorylated (p-JNK), procaspase-3, BCL2-associated X protein (Bax), Bcl-2 (B-cell lymphoma 2), and poly [ADP-ribose] polymerase 1 (PARP-1) proteins expression levels with or without JNK inhibitor (SP600125) in the HT22 cell line. The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. Number of experiments performed N = 3. b , c Immunofluorescence images of activated p-JNK (green) and caspase-3 (red) proteins along with their relative histograms after drug treatment with d -gal (100 mM), Gly (20 μg/μl), and SP600125 (20 μM) treatment in HT22 cell line for 24 h. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. The data are expressed as the mean ± SEM. Magnification × 40. Scale bar; 50 μm. a Significantly different from the control group while bcd significantly different from the d -gal-treated groups. Significance: a, b, c, d P

    Journal: Journal of Neuroinflammation

    Article Title: Glycine, the smallest amino acid, confers neuroprotection against d-galactose-induced neurodegeneration and memory impairment by regulating c-Jun N-terminal kinase in the mouse brain

    doi: 10.1186/s12974-020-01989-w

    Figure Lengend Snippet: Glycine treatment reduced d -galactose-mediated elevated p-JNK-dependent neuroapoptosis in HT22 cells lines. a Representative western blot analysis of activated phosphorylated (p-JNK), procaspase-3, BCL2-associated X protein (Bax), Bcl-2 (B-cell lymphoma 2), and poly [ADP-ribose] polymerase 1 (PARP-1) proteins expression levels with or without JNK inhibitor (SP600125) in the HT22 cell line. The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. Number of experiments performed N = 3. b , c Immunofluorescence images of activated p-JNK (green) and caspase-3 (red) proteins along with their relative histograms after drug treatment with d -gal (100 mM), Gly (20 μg/μl), and SP600125 (20 μM) treatment in HT22 cell line for 24 h. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. The data are expressed as the mean ± SEM. Magnification × 40. Scale bar; 50 μm. a Significantly different from the control group while bcd significantly different from the d -gal-treated groups. Significance: a, b, c, d P

    Article Snippet: To confirm equal sample loading, an anti-β-actin antibody (Santa Cruz Biotechnology, Dallas, TX, USA) was used as a standard for comparison.

    Techniques: Western Blot, Expressing, Software, Immunofluorescence, Staining

    Glycine treatment inhibited d -galactose-induced oxidative stress and ameliorates ROS/LPO production in mice brain. a , b Representative histograms of reactive oxygen species (ROS measured as DCF level) and lipid peroxidation (LPO measured as MDA level) assays of Gly against d -galactose in the cortex and hippocampus of mice brain. c – e Western blot analysis representing the expression level of Nrf2 and HO1 proteins in the hippocampus of the experimental groups. The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. g The immunofluorescence images represented the immunoreactivity of Nrf2 proteins (Red, TRITC; Blue, DAPI) in the cortex and hippocampus regions of mice brain along with their relative histograms, respectively. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions =100 μm. Asterisk (*) sign indicated significant difference from the normal saline treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated control group. Significance: * P ≤ 0.05, ** P ≤ 0.01; *** P ≤ 0.001; # P ≤ 0.05, ## P ≤ 0.01

    Journal: Journal of Neuroinflammation

    Article Title: Glycine, the smallest amino acid, confers neuroprotection against d-galactose-induced neurodegeneration and memory impairment by regulating c-Jun N-terminal kinase in the mouse brain

    doi: 10.1186/s12974-020-01989-w

    Figure Lengend Snippet: Glycine treatment inhibited d -galactose-induced oxidative stress and ameliorates ROS/LPO production in mice brain. a , b Representative histograms of reactive oxygen species (ROS measured as DCF level) and lipid peroxidation (LPO measured as MDA level) assays of Gly against d -galactose in the cortex and hippocampus of mice brain. c – e Western blot analysis representing the expression level of Nrf2 and HO1 proteins in the hippocampus of the experimental groups. The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. g The immunofluorescence images represented the immunoreactivity of Nrf2 proteins (Red, TRITC; Blue, DAPI) in the cortex and hippocampus regions of mice brain along with their relative histograms, respectively. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions =100 μm. Asterisk (*) sign indicated significant difference from the normal saline treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated control group. Significance: * P ≤ 0.05, ** P ≤ 0.01; *** P ≤ 0.001; # P ≤ 0.05, ## P ≤ 0.01

    Article Snippet: To confirm equal sample loading, an anti-β-actin antibody (Santa Cruz Biotechnology, Dallas, TX, USA) was used as a standard for comparison.

    Techniques: Mouse Assay, Multiple Displacement Amplification, Western Blot, Expressing, Software, Immunofluorescence, Staining

    Gly inhibited d -galactose-induced activation of inflammatory proteins in the hippocampus of mice brain. a The Western blot analysis of tumor necrosis factor alpha (TNF-α), interleukin-1 βeta (IL-1βeta), glial fibrillary acidic protein (GFAP), and ionized calcium binding adaptor molecule 1 (Iba1) protein expression level in the hippocampus of mice The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. b , c The immunofluorescence images represent the immunoreactivity of IL-1βeta (green, FITC; Blue, DAPI) in cortex and hippocampus of mice ( d ) The immunofluorescence images represent the immunoreactivity of GFAP (green, FITC; blue, DAPI) in cortex and hippocampus (CA-1 and DG regions) of mice The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions =100 μm. Asterisk (*) sign indicated significant difference from the normal saline-treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated control group. Significance: * P ≤ 0.05, ** P ≤ 0.01; *** P ≤ 0.001; # P ≤ 0.05, ## P ≤ 0.01.

    Journal: Journal of Neuroinflammation

    Article Title: Glycine, the smallest amino acid, confers neuroprotection against d-galactose-induced neurodegeneration and memory impairment by regulating c-Jun N-terminal kinase in the mouse brain

    doi: 10.1186/s12974-020-01989-w

    Figure Lengend Snippet: Gly inhibited d -galactose-induced activation of inflammatory proteins in the hippocampus of mice brain. a The Western blot analysis of tumor necrosis factor alpha (TNF-α), interleukin-1 βeta (IL-1βeta), glial fibrillary acidic protein (GFAP), and ionized calcium binding adaptor molecule 1 (Iba1) protein expression level in the hippocampus of mice The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. b , c The immunofluorescence images represent the immunoreactivity of IL-1βeta (green, FITC; Blue, DAPI) in cortex and hippocampus of mice ( d ) The immunofluorescence images represent the immunoreactivity of GFAP (green, FITC; blue, DAPI) in cortex and hippocampus (CA-1 and DG regions) of mice The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions =100 μm. Asterisk (*) sign indicated significant difference from the normal saline-treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated control group. Significance: * P ≤ 0.05, ** P ≤ 0.01; *** P ≤ 0.001; # P ≤ 0.05, ## P ≤ 0.01.

    Article Snippet: To confirm equal sample loading, an anti-β-actin antibody (Santa Cruz Biotechnology, Dallas, TX, USA) was used as a standard for comparison.

    Techniques: Activation Assay, Mouse Assay, Western Blot, Binding Assay, Expressing, Software, Immunofluorescence, Staining

    Glycine treatment inhibited d -galactose-induced elevated p-JNK and apoptotic cell death in mice brain. a Representative western blot analysis of stress kinase phosphorylated (p-JNK), cleaved caspase-3, cytochrome c (Cyt. C), Bcl-2 (B-cell lymphoma 2), and PARP-1 (poly-ADP-ribosyltransferase) proteins expression levels in both cortex and hippocampus regions of mice brain. The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. b Representative immunofluorescence results of caspase-3 (red; in hippocampus) and activated p-JNK proteins (green; in cortex and hippocampus) of the experimental mice group and c , d Nissl (cortex, DG, CA1, and CA3 regions) FJB staining (green, FITC; Blue) in cortex and hippocampus regions of experimental mice brain. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions =100 μm. Asterisk (*) sign indicated significant difference from the normal saline treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated control group. Significance: * P ≤ 0.05, ** P ≤ 0.01; *** P ≤ 0.001; # P ≤ 0.05, ## P ≤ 0.01

    Journal: Journal of Neuroinflammation

    Article Title: Glycine, the smallest amino acid, confers neuroprotection against d-galactose-induced neurodegeneration and memory impairment by regulating c-Jun N-terminal kinase in the mouse brain

    doi: 10.1186/s12974-020-01989-w

    Figure Lengend Snippet: Glycine treatment inhibited d -galactose-induced elevated p-JNK and apoptotic cell death in mice brain. a Representative western blot analysis of stress kinase phosphorylated (p-JNK), cleaved caspase-3, cytochrome c (Cyt. C), Bcl-2 (B-cell lymphoma 2), and PARP-1 (poly-ADP-ribosyltransferase) proteins expression levels in both cortex and hippocampus regions of mice brain. The cropped bands were quantified using ImageJ software, and the differences are represented in the histogram. The density values are expressed in arbitrary units (A.U.) as the mean ± SEM for the respective indicated protein. An anti-β-actin antibody was used as a loading control. n = 8 mice/group, and the number of experiments performed N = 3. b Representative immunofluorescence results of caspase-3 (red; in hippocampus) and activated p-JNK proteins (green; in cortex and hippocampus) of the experimental mice group and c , d Nissl (cortex, DG, CA1, and CA3 regions) FJB staining (green, FITC; Blue) in cortex and hippocampus regions of experimental mice brain. The relative integrated density values are represented in arbitrary units (A.U) as the means (± S.E.M) for the respective indicated proteins. DAPI (blue) was used for nucleus staining. n = 8 mice/group, and the number of experiments = 3. Magnification × 40. Scale bar; 50 μm = cortices; DG hippocampal regions =100 μm. Asterisk (*) sign indicated significant difference from the normal saline treated group; hash (#) sign indicated significant difference from d -gal-treated group; while the phi (Φ) sign indicated no significance from normal saline-treated control group. Significance: * P ≤ 0.05, ** P ≤ 0.01; *** P ≤ 0.001; # P ≤ 0.05, ## P ≤ 0.01

    Article Snippet: To confirm equal sample loading, an anti-β-actin antibody (Santa Cruz Biotechnology, Dallas, TX, USA) was used as a standard for comparison.

    Techniques: Mouse Assay, Western Blot, Expressing, Software, Immunofluorescence, Staining

    Monocyte chemoattractant protein‐1 mediates tumor growth factor β1‐primed fibrogenic activation of hepatic stellate cells. (A) Experimental approach to analyze the fibrogenic activation of hSCs. LX‐2 cells were treated with TGF‐ß1 for 1 hour and then supplied with fresh medium. MCP‐1 concentration in the culture supernatant was assessed at the indicated times. During cell cultivation, the culture medium was changed daily with or without the presence of MCP‐1 blocking antibody. At the end of the culturing phase, the cell count and type IV collagen protein levels were analyzed. (B) Assessment of extracellular secretion of MCP‐1 in the culture supernatant. Data represent mean ± SD. (C) Cell growth at 120 hours after transient TGF‐β1 stimulation (0, 10 ng/mL and 100 ng/mL) in the presence of either IgG control antibody or MCP‐1 blocking antibody ( # P

    Journal: Hepatology Communications

    Article Title: Role of monocyte chemoattractant protein‐1 in liver fibrosis with transient myeloproliferative disorder in down syndrome

    doi: 10.1002/hep4.1150

    Figure Lengend Snippet: Monocyte chemoattractant protein‐1 mediates tumor growth factor β1‐primed fibrogenic activation of hepatic stellate cells. (A) Experimental approach to analyze the fibrogenic activation of hSCs. LX‐2 cells were treated with TGF‐ß1 for 1 hour and then supplied with fresh medium. MCP‐1 concentration in the culture supernatant was assessed at the indicated times. During cell cultivation, the culture medium was changed daily with or without the presence of MCP‐1 blocking antibody. At the end of the culturing phase, the cell count and type IV collagen protein levels were analyzed. (B) Assessment of extracellular secretion of MCP‐1 in the culture supernatant. Data represent mean ± SD. (C) Cell growth at 120 hours after transient TGF‐β1 stimulation (0, 10 ng/mL and 100 ng/mL) in the presence of either IgG control antibody or MCP‐1 blocking antibody ( # P

    Article Snippet: Cells were stimulated with either recombinant human MCP‐1 (Z028029, GenScript, Piscataway, NJ) or TGF‐β1 (240‐B‐002, R & D Systems, Minneapolis, MN) and then incubated with either 1 μg/mL MCP‐1 blocking antibody (M2420; Sigma, St. Louis, MO) or mouse IgG (278‐810; Ancell, Bayport, MN).

    Techniques: Activation Assay, Concentration Assay, Blocking Assay, Cell Counting

    Liver, 22-month-old male C57BL/6J mouse. Hepatoblastoma (left) and adjacent hepatocellular adenoma (right). β-catenin immunohistochemistry reveals a strong nucleocytoplasmic signal in hepatoblastoma tumor cells, while neoplastic hepatocytes in the surrounding adenoma display a delicate membranous staining pattern. β-catenin immunohistochemistry (IHC); hematoxylin counterstain. Scale bar: 100 µm.

    Journal: Journal of Toxicologic Pathology

    Article Title: Spontaneous pulmonary co-metastasis of hepatoblastoma arising within a hepatocellular carcinoma in an aged C57BL/6J mouse

    doi: 10.1293/tox.2017-0067

    Figure Lengend Snippet: Liver, 22-month-old male C57BL/6J mouse. Hepatoblastoma (left) and adjacent hepatocellular adenoma (right). β-catenin immunohistochemistry reveals a strong nucleocytoplasmic signal in hepatoblastoma tumor cells, while neoplastic hepatocytes in the surrounding adenoma display a delicate membranous staining pattern. β-catenin immunohistochemistry (IHC); hematoxylin counterstain. Scale bar: 100 µm.

    Article Snippet: Endogenous peroxidase was blocked by incubating sections in 3% H2 O2 for 15 min. For antigen retrieval, sections were immersed in citrate buffer (pH 6.0), heated, and cooled at room temperature. β-catenin (rabbit monoclonal, clone 6B3, dilution 1:150, Cell Signaling) specific antibody was applied for 1.5 hours at room temperature.

    Techniques: Immunohistochemistry, Staining

    Repeated Plasmodium vivax exposures increase the levels of antigen-specific IgG. A-C. Components of humoral response were measured in patients acutely infected for the first time (black squares) or with multiple infections (blue squares) with P . vivax . A. Pv AMA-1 and PvMSP-1 19 IgM and IgG were measure in plasma. B. Scattered plots showing frequency of activated memory (CD27 + CD21 - ), classical memory (CD27 + CD21 + ), naïve (CD27 - CD21 + ) and atypical memory (CD27 - CD21 - ) B cells in P . vivax -infected patients described above. C. Scattered plots showing the proportion of plasma cells (CD21 - CD20 - ) and IgG, CD38, Ki67, PD-1, IgG and PD-1 and Ki67 and CD38 expressing plasma cells from P . vivax -infected patients. All the B cell subsets were analyzed after gating on live CD19 + cells. D. Frequency of Tfh cells (PD-1 + ICOS + CXCR5 + CD45RO + CD4 + CD3 + ) cells are shown in patients infected for the first time (black squares) or infected 2 to 5 times (grey squares) or more than 5 times (blue squares) with P . vivax . Lines represent median values of the given measurement in each group (left graph). Correlation between number of malaria episodes and proportion of Tfh cells (right graph). p values are depicted in the figure.

    Journal: PLoS Pathogens

    Article Title: T follicular helper cells regulate the activation of B lymphocytes and antibody production during Plasmodium vivax infection

    doi: 10.1371/journal.ppat.1006484

    Figure Lengend Snippet: Repeated Plasmodium vivax exposures increase the levels of antigen-specific IgG. A-C. Components of humoral response were measured in patients acutely infected for the first time (black squares) or with multiple infections (blue squares) with P . vivax . A. Pv AMA-1 and PvMSP-1 19 IgM and IgG were measure in plasma. B. Scattered plots showing frequency of activated memory (CD27 + CD21 - ), classical memory (CD27 + CD21 + ), naïve (CD27 - CD21 + ) and atypical memory (CD27 - CD21 - ) B cells in P . vivax -infected patients described above. C. Scattered plots showing the proportion of plasma cells (CD21 - CD20 - ) and IgG, CD38, Ki67, PD-1, IgG and PD-1 and Ki67 and CD38 expressing plasma cells from P . vivax -infected patients. All the B cell subsets were analyzed after gating on live CD19 + cells. D. Frequency of Tfh cells (PD-1 + ICOS + CXCR5 + CD45RO + CD4 + CD3 + ) cells are shown in patients infected for the first time (black squares) or infected 2 to 5 times (grey squares) or more than 5 times (blue squares) with P . vivax . Lines represent median values of the given measurement in each group (left graph). Correlation between number of malaria episodes and proportion of Tfh cells (right graph). p values are depicted in the figure.

    Article Snippet: The presence of bound IgA, IgM, IgG and subclasses of IgG was detected using tetramethylbenzidine (Sigma) at 10mg/mL diluted in phosphate-citrate buffer (pH 5.0) containing hydrogen peroxide (0.03% [vol/vol]).

    Techniques: Infection, Expressing

    Increase in the reactivity index of IgM and IgG against MSP-1 19 from Plasmodium vivax during malaria. A. Total and Plasmodium vivax -specific antibodies were measured by enzyme-linked immunosorbent assay (ELISA). Total and PvMSP-1 19 IgM and IgG were measured in plasma of patients during acute malaria (BT) and after treatment (AT). B. IgG subclasses against PvMSP-1 19 were measure in plasma of patients during acute malaria (BT) and after treatment (AT). Lines represent median values of the given measurement in each group. Dotted lines represent healthy donors. p values are depicted in the figure.

    Journal: PLoS Pathogens

    Article Title: T follicular helper cells regulate the activation of B lymphocytes and antibody production during Plasmodium vivax infection

    doi: 10.1371/journal.ppat.1006484

    Figure Lengend Snippet: Increase in the reactivity index of IgM and IgG against MSP-1 19 from Plasmodium vivax during malaria. A. Total and Plasmodium vivax -specific antibodies were measured by enzyme-linked immunosorbent assay (ELISA). Total and PvMSP-1 19 IgM and IgG were measured in plasma of patients during acute malaria (BT) and after treatment (AT). B. IgG subclasses against PvMSP-1 19 were measure in plasma of patients during acute malaria (BT) and after treatment (AT). Lines represent median values of the given measurement in each group. Dotted lines represent healthy donors. p values are depicted in the figure.

    Article Snippet: The presence of bound IgA, IgM, IgG and subclasses of IgG was detected using tetramethylbenzidine (Sigma) at 10mg/mL diluted in phosphate-citrate buffer (pH 5.0) containing hydrogen peroxide (0.03% [vol/vol]).

    Techniques: Enzyme-linked Immunosorbent Assay

    Immunohistochemical analysis of β-catenin and c-Myc protein expression in the venous limb of the murine AVF 1 wk after the creation of the AVF. Immunohistochemical localization of β-catenin protein in control vein ( A ) and the venous segment

    Journal: American Journal of Physiology - Renal Physiology

    Article Title: ?-Catenin is markedly induced in a murine model of an arteriovenous fistula: the effect of metalloproteinase inhibition

    doi: 10.1152/ajprenal.00488.2010

    Figure Lengend Snippet: Immunohistochemical analysis of β-catenin and c-Myc protein expression in the venous limb of the murine AVF 1 wk after the creation of the AVF. Immunohistochemical localization of β-catenin protein in control vein ( A ) and the venous segment

    Article Snippet: Primary rabbit polyclonal antibodies to c-Myc and β-catenin (catalog nos. sc-764 and sc-1496R, respectively; Santa Cruz Biotechnology, Santa Cruz, CA) were applied for 60 min at room temperature.

    Techniques: Immunohistochemistry, Expressing

    The effect of doxycycline on N-cadherin, β-catenin, and c-Myc protein expression in the venous limb of the murine AVF 1 wk after the creation of the AVF. Protein extracted from the venous segment of the AVF in mice with or without doxycycline

    Journal: American Journal of Physiology - Renal Physiology

    Article Title: ?-Catenin is markedly induced in a murine model of an arteriovenous fistula: the effect of metalloproteinase inhibition

    doi: 10.1152/ajprenal.00488.2010

    Figure Lengend Snippet: The effect of doxycycline on N-cadherin, β-catenin, and c-Myc protein expression in the venous limb of the murine AVF 1 wk after the creation of the AVF. Protein extracted from the venous segment of the AVF in mice with or without doxycycline

    Article Snippet: Primary rabbit polyclonal antibodies to c-Myc and β-catenin (catalog nos. sc-764 and sc-1496R, respectively; Santa Cruz Biotechnology, Santa Cruz, CA) were applied for 60 min at room temperature.

    Techniques: Expressing, Mouse Assay

    Western analysis of β-catenin protein expression in the venous limb of the murine AVF 1 wk after the creation of the AVF. Protein extracted from control and AVF veins was immunoblotted for β-catenin. Equivalency of protein loading was

    Journal: American Journal of Physiology - Renal Physiology

    Article Title: ?-Catenin is markedly induced in a murine model of an arteriovenous fistula: the effect of metalloproteinase inhibition

    doi: 10.1152/ajprenal.00488.2010

    Figure Lengend Snippet: Western analysis of β-catenin protein expression in the venous limb of the murine AVF 1 wk after the creation of the AVF. Protein extracted from control and AVF veins was immunoblotted for β-catenin. Equivalency of protein loading was

    Article Snippet: Primary rabbit polyclonal antibodies to c-Myc and β-catenin (catalog nos. sc-764 and sc-1496R, respectively; Santa Cruz Biotechnology, Santa Cruz, CA) were applied for 60 min at room temperature.

    Techniques: Western Blot, Expressing