lysophosphatidic acid lpa  (Echelon Biosciences)


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    Echelon Biosciences lysophosphatidic acid lpa
    Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; <t>LPA,</t> <t>lysophosphatidic</t> acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.
    Lysophosphatidic Acid Lpa, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/lysophosphatidic acid lpa/product/Echelon Biosciences
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    lysophosphatidic acid lpa - by Bioz Stars, 2023-02
    93/100 stars

    Images

    1) Product Images from "Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins"

    Article Title: Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

    Journal: Molecular & Cellular Proteomics : MCP

    doi: 10.1016/j.mcpro.2021.100102

    Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.
    Figure Legend Snippet: Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.

    Techniques Used: Stripping Membranes, Recombinant, Binding Assay, Mutagenesis, Confocal Microscopy, Staining, Incubation

    PARP1 binds to PPIn via three polybasic regions. A , domain structure of PARP1 and deletion constructs. B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank. C and D , lipid overlay assay using PIP strips incubated with recombinant GST-PARP1 deletion constructs WT and mutants and detection of protein–lipid interactions using an anti-GST-HRP–conjugated antibody. E , multiple sequence alignment of the polybasic regions located in the zinc finger III and the BRCT-WGR linker found in human PARP-1 compared with other vertebrate species performed using the online program MUSCLE. Accession number for Homo sapiens (P09874), Mus musculus (P11103), Bos taurus (P18493), Gallus gallus (P26446), Xenopus laevis (P31669), and Danio rerio (Q5RHR0). F , ribbon representation of the human PARP-1 zinc-finger III NMR structure (aa 233–357 PDB: 2JVN, ). The two polybasic regions found in the N-terminal (aa 233–236) and C-terminal (aa 346–352) parts of the zinc-finger III are colored in red . ART, (ADP-ribosyl) transferase domain (ART); BRCT, BRCA1 C-terminal domain; HD, helical subdomain; HRP, horse radish peroxidase; PARP1, poly(ADP-ribose) polymerase 1; ZnF 1 to 3, zinc-finger I-III; WGR, Trp-Gly-Arg.
    Figure Legend Snippet: PARP1 binds to PPIn via three polybasic regions. A , domain structure of PARP1 and deletion constructs. B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank. C and D , lipid overlay assay using PIP strips incubated with recombinant GST-PARP1 deletion constructs WT and mutants and detection of protein–lipid interactions using an anti-GST-HRP–conjugated antibody. E , multiple sequence alignment of the polybasic regions located in the zinc finger III and the BRCT-WGR linker found in human PARP-1 compared with other vertebrate species performed using the online program MUSCLE. Accession number for Homo sapiens (P09874), Mus musculus (P11103), Bos taurus (P18493), Gallus gallus (P26446), Xenopus laevis (P31669), and Danio rerio (Q5RHR0). F , ribbon representation of the human PARP-1 zinc-finger III NMR structure (aa 233–357 PDB: 2JVN, ). The two polybasic regions found in the N-terminal (aa 233–236) and C-terminal (aa 346–352) parts of the zinc-finger III are colored in red . ART, (ADP-ribosyl) transferase domain (ART); BRCT, BRCA1 C-terminal domain; HD, helical subdomain; HRP, horse radish peroxidase; PARP1, poly(ADP-ribose) polymerase 1; ZnF 1 to 3, zinc-finger I-III; WGR, Trp-Gly-Arg.

    Techniques Used: Construct, Overlay Assay, Incubation, Recombinant, Sequencing

    lysophosphatidic acid lpa  (Echelon Biosciences)


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    Echelon Biosciences lysophosphatidic acid lpa
    Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; <t>LPA,</t> <t>lysophosphatidic</t> acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.
    Lysophosphatidic Acid Lpa, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/lysophosphatidic acid lpa/product/Echelon Biosciences
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    lysophosphatidic acid lpa - by Bioz Stars, 2023-02
    93/100 stars

    Images

    1) Product Images from "Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins"

    Article Title: Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

    Journal: Molecular & Cellular Proteomics : MCP

    doi: 10.1016/j.mcpro.2021.100102

    Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.
    Figure Legend Snippet: Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.

    Techniques Used: Stripping Membranes, Recombinant, Binding Assay, Mutagenesis, Confocal Microscopy, Staining, Incubation

    PARP1 binds to PPIn via three polybasic regions. A , domain structure of PARP1 and deletion constructs. B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank. C and D , lipid overlay assay using PIP strips incubated with recombinant GST-PARP1 deletion constructs WT and mutants and detection of protein–lipid interactions using an anti-GST-HRP–conjugated antibody. E , multiple sequence alignment of the polybasic regions located in the zinc finger III and the BRCT-WGR linker found in human PARP-1 compared with other vertebrate species performed using the online program MUSCLE. Accession number for Homo sapiens (P09874), Mus musculus (P11103), Bos taurus (P18493), Gallus gallus (P26446), Xenopus laevis (P31669), and Danio rerio (Q5RHR0). F , ribbon representation of the human PARP-1 zinc-finger III NMR structure (aa 233–357 PDB: 2JVN, ). The two polybasic regions found in the N-terminal (aa 233–236) and C-terminal (aa 346–352) parts of the zinc-finger III are colored in red . ART, (ADP-ribosyl) transferase domain (ART); BRCT, BRCA1 C-terminal domain; HD, helical subdomain; HRP, horse radish peroxidase; PARP1, poly(ADP-ribose) polymerase 1; ZnF 1 to 3, zinc-finger I-III; WGR, Trp-Gly-Arg.
    Figure Legend Snippet: PARP1 binds to PPIn via three polybasic regions. A , domain structure of PARP1 and deletion constructs. B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank. C and D , lipid overlay assay using PIP strips incubated with recombinant GST-PARP1 deletion constructs WT and mutants and detection of protein–lipid interactions using an anti-GST-HRP–conjugated antibody. E , multiple sequence alignment of the polybasic regions located in the zinc finger III and the BRCT-WGR linker found in human PARP-1 compared with other vertebrate species performed using the online program MUSCLE. Accession number for Homo sapiens (P09874), Mus musculus (P11103), Bos taurus (P18493), Gallus gallus (P26446), Xenopus laevis (P31669), and Danio rerio (Q5RHR0). F , ribbon representation of the human PARP-1 zinc-finger III NMR structure (aa 233–357 PDB: 2JVN, ). The two polybasic regions found in the N-terminal (aa 233–236) and C-terminal (aa 346–352) parts of the zinc-finger III are colored in red . ART, (ADP-ribosyl) transferase domain (ART); BRCT, BRCA1 C-terminal domain; HD, helical subdomain; HRP, horse radish peroxidase; PARP1, poly(ADP-ribose) polymerase 1; ZnF 1 to 3, zinc-finger I-III; WGR, Trp-Gly-Arg.

    Techniques Used: Construct, Overlay Assay, Incubation, Recombinant, Sequencing

    human lpa elisa kit  (Echelon Biosciences)


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    Echelon Biosciences human lpa elisa kit
    Effect of lysophosphatidic acid <t>(LPA)</t> on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by <t>ELISA.</t> Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001
    Human Lpa Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/human lpa elisa kit/product/Echelon Biosciences
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    human lpa elisa kit - by Bioz Stars, 2023-02
    93/100 stars

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    1) Product Images from "CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts"

    Article Title: CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts

    Journal: Respiratory Research

    doi: 10.1186/s12931-021-01677-0

    Effect of lysophosphatidic acid (LPA) on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001
    Figure Legend Snippet: Effect of lysophosphatidic acid (LPA) on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001

    Techniques Used: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of CREB inhibitor (CREB inh) on growth factor mRNA and protein levels. Primary basal cells from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 200 nM CREB inhibitor 666-15 and a combination of LPA and CREB inhibitor 666-15. a mRNA expression levels. Shown are connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) mRNA levels determined after 3 h of LPA exposure with and without CREB inhibitor (666-15) using qRT-PCR using 18S RNA to normalize the samples. b Protein levels. Shown are protein levels in conditioned media of LPA with and without CREB inhibitor 666-15, LPA with or without CREB inhibitor. After 3 h, the stimuli/inhibitor were removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor levels by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND not detected. *p < 0.05, **p < 0.01, ***p < 0.001
    Figure Legend Snippet: Effect of CREB inhibitor (CREB inh) on growth factor mRNA and protein levels. Primary basal cells from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 200 nM CREB inhibitor 666-15 and a combination of LPA and CREB inhibitor 666-15. a mRNA expression levels. Shown are connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) mRNA levels determined after 3 h of LPA exposure with and without CREB inhibitor (666-15) using qRT-PCR using 18S RNA to normalize the samples. b Protein levels. Shown are protein levels in conditioned media of LPA with and without CREB inhibitor 666-15, LPA with or without CREB inhibitor. After 3 h, the stimuli/inhibitor were removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor levels by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Techniques Used: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of ERK1/2 inhibitor (ERK1/2 inh) on growth factor expression. Primary basal cells from each of 3 non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 5 µM ERK1/2 inhibitor (LY32149966) and a combination of LPA and ERK1/2 inhibitor (LY32149966). a mRNA. Shown are expression levels of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) determined after 3 h of LPA exposure with and without ERK1/2 inhibitor (LY32149966) using qRT-PCR using 18S RNA to normalize the samples. b Protein. Shown are growth factor levels in the conditioned media. After 3 h of LPA with and without CREB inhibitor (666-15) exposure, LPA with or without ERK1/2 inhibitor was removed and cells were incubated with un-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001
    Figure Legend Snippet: Effect of ERK1/2 inhibitor (ERK1/2 inh) on growth factor expression. Primary basal cells from each of 3 non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 5 µM ERK1/2 inhibitor (LY32149966) and a combination of LPA and ERK1/2 inhibitor (LY32149966). a mRNA. Shown are expression levels of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) determined after 3 h of LPA exposure with and without ERK1/2 inhibitor (LY32149966) using qRT-PCR using 18S RNA to normalize the samples. b Protein. Shown are growth factor levels in the conditioned media. After 3 h of LPA with and without CREB inhibitor (666-15) exposure, LPA with or without ERK1/2 inhibitor was removed and cells were incubated with un-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Techniques Used: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of EGFR inhibitor (EGFR inh) on growth factor mRNA levels. Primary basal cells (passage 3) from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 10 µM EGFR inhibitor (AG1478), and a combination of LPA and EGFR inhibitor. a mRNA expression levels shown are mRNA levels of connective tissue growth factor ( CTGF ), endothelin-1( EDN1 ), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor B were determined after 3 h of LPA exposure with and without the inhibitor using qRT-PCR using 18S RNA to normalize the samples. b Protein levels in the conditioned media. After 3 h of LPA with and without EGFR inhibitor (AG1478) exposure, LPA with or without inhibitor was removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001
    Figure Legend Snippet: Effect of EGFR inhibitor (EGFR inh) on growth factor mRNA levels. Primary basal cells (passage 3) from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 10 µM EGFR inhibitor (AG1478), and a combination of LPA and EGFR inhibitor. a mRNA expression levels shown are mRNA levels of connective tissue growth factor ( CTGF ), endothelin-1( EDN1 ), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor B were determined after 3 h of LPA exposure with and without the inhibitor using qRT-PCR using 18S RNA to normalize the samples. b Protein levels in the conditioned media. After 3 h of LPA with and without EGFR inhibitor (AG1478) exposure, LPA with or without inhibitor was removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Techniques Used: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of CREB inhibitor (CREB inh) on fibroblast expression in response to basal cell conditioned medium. Primary normal human lung fibroblasts (NHLF) were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Fibroblast expression of collagen type 1 ( COL1A1 ) and smooth muscle actin ( ACTA2 ) was assessed after 24 h in conditioned medium. a mRNA levels. RNA was harvested and evaluated by qRT-PCR using 18S RNA to normalize the samples. b Protein levels. At the same time point, conditioned media was collected and assessed for COL1A1 secretion. Cells were solubilized with a RIPA buffer and evaluated for alpha smooth muscle myosin by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001
    Figure Legend Snippet: Effect of CREB inhibitor (CREB inh) on fibroblast expression in response to basal cell conditioned medium. Primary normal human lung fibroblasts (NHLF) were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Fibroblast expression of collagen type 1 ( COL1A1 ) and smooth muscle actin ( ACTA2 ) was assessed after 24 h in conditioned medium. a mRNA levels. RNA was harvested and evaluated by qRT-PCR using 18S RNA to normalize the samples. b Protein levels. At the same time point, conditioned media was collected and assessed for COL1A1 secretion. Cells were solubilized with a RIPA buffer and evaluated for alpha smooth muscle myosin by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001

    Techniques Used: Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

    Effect of CREB inhibitor (CREB inh) on autotaxin ( ENPP2 ) expression, secretion, and activity in fibroblasts in response to medium conditioned by LPA-treated basal cells. Primary NHLF were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1.0 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Autotaxin, an enzyme responsible for the extracellular production of LPA, in encoded by ENPP2 . Expression of the ENPP2 gene was assessed in fibroblasts 24 h after exposure to conditioned medium. Autotaxin protein levels and LPA levels were assessed in cell culture medium 24 h after exposure to conditioned medium. a ENPP2 mRNA levels. mRNA was measured by qRT-PCR using 18S RNA to normalize the samples. b Autotaxin protein levels. Protein levels were measured in cell culture medium by ELISA. c LPA levels. LPA was measured in cell culture medium by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001
    Figure Legend Snippet: Effect of CREB inhibitor (CREB inh) on autotaxin ( ENPP2 ) expression, secretion, and activity in fibroblasts in response to medium conditioned by LPA-treated basal cells. Primary NHLF were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1.0 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Autotaxin, an enzyme responsible for the extracellular production of LPA, in encoded by ENPP2 . Expression of the ENPP2 gene was assessed in fibroblasts 24 h after exposure to conditioned medium. Autotaxin protein levels and LPA levels were assessed in cell culture medium 24 h after exposure to conditioned medium. a ENPP2 mRNA levels. mRNA was measured by qRT-PCR using 18S RNA to normalize the samples. b Autotaxin protein levels. Protein levels were measured in cell culture medium by ELISA. c LPA levels. LPA was measured in cell culture medium by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001

    Techniques Used: Expressing, Activity Assay, Cell Culture, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

    lysophosphatidic acid assay kit  (Echelon Biosciences)


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    Echelon Biosciences lysophosphatidic acid assay kit
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    lpa assay kit  (Echelon Biosciences)


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    Echelon Biosciences lpa assay kit
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    Echelon Biosciences lpa assay kit
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    lysophosphatidic acid  (Echelon Biosciences)


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    Echelon Biosciences lysophosphatidic acid assay kit ii
    Experimental procedures used for determination of endometrial LPA, LPAR1–4 mRNA level and protein abundance, effect of LPA on PG and CTGF secretion at different stages of endometrosis. Ctr-control, <t> LPA-lysophosphatidic </t> acid, AA- arachidonic acid
    Lysophosphatidic Acid Assay Kit Ii, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Lysophosphatidic acid as a regulator of endometrial connective tissue growth factor and prostaglandin secretion during estrous cycle and endometrosis in the mare"

    Article Title: Lysophosphatidic acid as a regulator of endometrial connective tissue growth factor and prostaglandin secretion during estrous cycle and endometrosis in the mare

    Journal: BMC Veterinary Research

    doi: 10.1186/s12917-020-02562-6

    Experimental procedures used for determination of endometrial LPA, LPAR1–4 mRNA level and protein abundance, effect of LPA on PG and CTGF secretion at different stages of endometrosis. Ctr-control,  LPA-lysophosphatidic  acid, AA- arachidonic acid
    Figure Legend Snippet: Experimental procedures used for determination of endometrial LPA, LPAR1–4 mRNA level and protein abundance, effect of LPA on PG and CTGF secretion at different stages of endometrosis. Ctr-control, LPA-lysophosphatidic acid, AA- arachidonic acid

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    lpa assay kit ii k 2800s  (Echelon Biosciences)


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    Echelon Biosciences lpa assay kit ii k 2800s
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    lysophosphatidic acid competitive elisa kit  (Echelon Biosciences)


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    Echelon Biosciences lysophosphatidic acid competitive elisa kit
    <t>Lysophosphatidic</t> acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.
    Lysophosphatidic Acid Competitive Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Protective Effect of Phosphatidylserine Blockade in Hemorrhagic Shock Phosphatidylserine Blockade in Hemorrhage"

    Article Title: Protective Effect of Phosphatidylserine Blockade in Hemorrhagic Shock Phosphatidylserine Blockade in Hemorrhage

    Journal: The Journal of surgical research

    doi: 10.1016/j.jss.2019.07.050

    Lysophosphatidic acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.
    Figure Legend Snippet: Lysophosphatidic acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.

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    lysophosphatidic acid competitive elisa kit  (Echelon Biosciences)


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    Echelon Biosciences lysophosphatidic acid competitive elisa kit
    <t>Lysophosphatidic</t> acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.
    Lysophosphatidic Acid Competitive Elisa Kit, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 93 stars, based on 1 article reviews
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    lysophosphatidic acid competitive elisa kit - by Bioz Stars, 2023-02
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    1) Product Images from "Protective Effect of Phosphatidylserine Blockade in Hemorrhagic Shock Phosphatidylserine Blockade in Hemorrhage"

    Article Title: Protective Effect of Phosphatidylserine Blockade in Hemorrhagic Shock Phosphatidylserine Blockade in Hemorrhage

    Journal: The Journal of surgical research

    doi: 10.1016/j.jss.2019.07.050

    Lysophosphatidic acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.
    Figure Legend Snippet: Lysophosphatidic acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.

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    Echelon Biosciences lysophosphatidic acid lpa
    Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; <t>LPA,</t> <t>lysophosphatidic</t> acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.
    Lysophosphatidic Acid Lpa, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Effect of lysophosphatidic acid <t>(LPA)</t> on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by <t>ELISA.</t> Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001
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    Echelon Biosciences lysophosphatidic acid assay kit
    Effect of lysophosphatidic acid <t>(LPA)</t> on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by <t>ELISA.</t> Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001
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    Echelon Biosciences lpa assay kit
    Effect of lysophosphatidic acid <t>(LPA)</t> on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by <t>ELISA.</t> Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001
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    Echelon Biosciences lysophosphatidic acid
    Effect of lysophosphatidic acid <t>(LPA)</t> on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by <t>ELISA.</t> Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001
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    Experimental procedures used for determination of endometrial LPA, LPAR1–4 mRNA level and protein abundance, effect of LPA on PG and CTGF secretion at different stages of endometrosis. Ctr-control, <t> LPA-lysophosphatidic </t> acid, AA- arachidonic acid
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    Experimental procedures used for determination of endometrial LPA, LPAR1–4 mRNA level and protein abundance, effect of LPA on PG and CTGF secretion at different stages of endometrosis. Ctr-control, <t> LPA-lysophosphatidic </t> acid, AA- arachidonic acid
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    Echelon Biosciences lysophosphatidic acid competitive elisa kit
    <t>Lysophosphatidic</t> acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.
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    Image Search Results


    Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.

    Journal: Molecular & Cellular Proteomics : MCP

    Article Title: Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

    doi: 10.1016/j.mcpro.2021.100102

    Figure Lengend Snippet: Specific detection of PtdIns(3,4,5) P 3 in the nucleolus. A , PIP strip (Echelon Inc) schematic showing the position of the spotted lipids each with 100 pmol. B , validation of the anti-PtdIns(3,4,5) P 3 antibody specificity using PIP strips. C , validation of the specificity of the recombinant GST-EGFP-GRP1-PH WT versus binding mutant K273A. D and E , confocal microscopy of actively growing HeLa cells stained with the indicated antibodies ( D ) or by incubation with recombinant GST-EGFP-GRP1-PH WT or K273A mutant combined with anti-nucleophosmin staining ( E ). F , quantification of the detection of nucleolar PtdIns(3,4,5) P 3 expressed as the percentage of HeLa cells showing foci detected by the GST-EGFP-GRP1-PH probe WT or K273A mutant within the area delimited by nucleophosmin (mean + SDs, n = 3, ∗ p < 0.05 two-way unpaired Student’s t test). The scale bar represents 5 μm. GRP1, general receptor for phosphoinositides-1; LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; PA, phosphatidic acid; PC, phosphatidylcholine; PE, phosphatidylethanolamine; PH, pleckstrin homology; PI, phosphatidylinositol; PIP3, PtdIns(3,4,5) P 3 ; PS, phosphatidylserine; PtdIns(3,4,5) P 3 , phosphatidylinositol 3,4,5-trisphosphate; S1P, sphingosine-1-phosphate; UBF, upstream binding factor.

    Article Snippet: B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank.

    Techniques: Stripping Membranes, Recombinant, Binding Assay, Mutagenesis, Confocal Microscopy, Staining, Incubation

    PARP1 binds to PPIn via three polybasic regions. A , domain structure of PARP1 and deletion constructs. B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank. C and D , lipid overlay assay using PIP strips incubated with recombinant GST-PARP1 deletion constructs WT and mutants and detection of protein–lipid interactions using an anti-GST-HRP–conjugated antibody. E , multiple sequence alignment of the polybasic regions located in the zinc finger III and the BRCT-WGR linker found in human PARP-1 compared with other vertebrate species performed using the online program MUSCLE. Accession number for Homo sapiens (P09874), Mus musculus (P11103), Bos taurus (P18493), Gallus gallus (P26446), Xenopus laevis (P31669), and Danio rerio (Q5RHR0). F , ribbon representation of the human PARP-1 zinc-finger III NMR structure (aa 233–357 PDB: 2JVN, ). The two polybasic regions found in the N-terminal (aa 233–236) and C-terminal (aa 346–352) parts of the zinc-finger III are colored in red . ART, (ADP-ribosyl) transferase domain (ART); BRCT, BRCA1 C-terminal domain; HD, helical subdomain; HRP, horse radish peroxidase; PARP1, poly(ADP-ribose) polymerase 1; ZnF 1 to 3, zinc-finger I-III; WGR, Trp-Gly-Arg.

    Journal: Molecular & Cellular Proteomics : MCP

    Article Title: Nuclear Phosphatidylinositol 3,4,5-Trisphosphate Interactome Uncovers an Enrichment in Nucleolar Proteins

    doi: 10.1016/j.mcpro.2021.100102

    Figure Lengend Snippet: PARP1 binds to PPIn via three polybasic regions. A , domain structure of PARP1 and deletion constructs. B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank. C and D , lipid overlay assay using PIP strips incubated with recombinant GST-PARP1 deletion constructs WT and mutants and detection of protein–lipid interactions using an anti-GST-HRP–conjugated antibody. E , multiple sequence alignment of the polybasic regions located in the zinc finger III and the BRCT-WGR linker found in human PARP-1 compared with other vertebrate species performed using the online program MUSCLE. Accession number for Homo sapiens (P09874), Mus musculus (P11103), Bos taurus (P18493), Gallus gallus (P26446), Xenopus laevis (P31669), and Danio rerio (Q5RHR0). F , ribbon representation of the human PARP-1 zinc-finger III NMR structure (aa 233–357 PDB: 2JVN, ). The two polybasic regions found in the N-terminal (aa 233–236) and C-terminal (aa 346–352) parts of the zinc-finger III are colored in red . ART, (ADP-ribosyl) transferase domain (ART); BRCT, BRCA1 C-terminal domain; HD, helical subdomain; HRP, horse radish peroxidase; PARP1, poly(ADP-ribose) polymerase 1; ZnF 1 to 3, zinc-finger I-III; WGR, Trp-Gly-Arg.

    Article Snippet: B , schematic representation of lipids spotted (100 pmol) on PIP strips (Echelon Biosciences) including lysophosphatidic acid (LPA), lysophosphatidylcholine (LPC), phosphatidylinositol (PtdIns), PtdIns3 P , PtdIns4 P , PtdIns5 P , phosphatidylethanolamine (PE), phosphatidylcholine (PC), sphingosine-1-Phosphate (S1P), PtdIns(3,4) P 2 , PtdIns(3,5) P 2 , PtdIns(4,5) P 2 , PtdIns(3,4,5) P 3 , phosphatic acid (PA), phosphatidylserine (PS), and blank.

    Techniques: Construct, Overlay Assay, Incubation, Recombinant, Sequencing

    Effect of lysophosphatidic acid (LPA) on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001

    Journal: Respiratory Research

    Article Title: CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts

    doi: 10.1186/s12931-021-01677-0

    Figure Lengend Snippet: Effect of lysophosphatidic acid (LPA) on fibrotic growth factor expression in the small airway epithelial basal cells (SAE BC). Primary SAE BC from each of 3 non-smoking individuals were plated in triplicate in the presence or absence of 1.0 μg/ml LPA and evaluated for expression of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 /ET-1), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor family member B ( PDGFB ). a mRNA. After a 3 h LPA exposure, RNA was harvested and evaluated by qRT-PCR normalized to 18S RNA. b Protein. After a 3 h LPA exposure, LPA was removed and cells were incubated with basal media without growth factor supplements for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Each sample was assessed in triplicate; data are expressed as the mean value of the 3 donors ± SE. *p < 0.05, **p < 0.01, ***p < 0.001

    Article Snippet: The human COL1A1 ELISA kit (MyBioSource, La Jolla, CA), ENPP2 human ELISA kit (R&D Systems), human LPA ELISA Kit (Echelon Biosciences) were performed according to the manufacturer’s instructions.

    Techniques: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of CREB inhibitor (CREB inh) on growth factor mRNA and protein levels. Primary basal cells from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 200 nM CREB inhibitor 666-15 and a combination of LPA and CREB inhibitor 666-15. a mRNA expression levels. Shown are connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) mRNA levels determined after 3 h of LPA exposure with and without CREB inhibitor (666-15) using qRT-PCR using 18S RNA to normalize the samples. b Protein levels. Shown are protein levels in conditioned media of LPA with and without CREB inhibitor 666-15, LPA with or without CREB inhibitor. After 3 h, the stimuli/inhibitor were removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor levels by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Journal: Respiratory Research

    Article Title: CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts

    doi: 10.1186/s12931-021-01677-0

    Figure Lengend Snippet: Effect of CREB inhibitor (CREB inh) on growth factor mRNA and protein levels. Primary basal cells from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 200 nM CREB inhibitor 666-15 and a combination of LPA and CREB inhibitor 666-15. a mRNA expression levels. Shown are connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) mRNA levels determined after 3 h of LPA exposure with and without CREB inhibitor (666-15) using qRT-PCR using 18S RNA to normalize the samples. b Protein levels. Shown are protein levels in conditioned media of LPA with and without CREB inhibitor 666-15, LPA with or without CREB inhibitor. After 3 h, the stimuli/inhibitor were removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor levels by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Article Snippet: The human COL1A1 ELISA kit (MyBioSource, La Jolla, CA), ENPP2 human ELISA kit (R&D Systems), human LPA ELISA Kit (Echelon Biosciences) were performed according to the manufacturer’s instructions.

    Techniques: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of ERK1/2 inhibitor (ERK1/2 inh) on growth factor expression. Primary basal cells from each of 3 non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 5 µM ERK1/2 inhibitor (LY32149966) and a combination of LPA and ERK1/2 inhibitor (LY32149966). a mRNA. Shown are expression levels of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) determined after 3 h of LPA exposure with and without ERK1/2 inhibitor (LY32149966) using qRT-PCR using 18S RNA to normalize the samples. b Protein. Shown are growth factor levels in the conditioned media. After 3 h of LPA with and without CREB inhibitor (666-15) exposure, LPA with or without ERK1/2 inhibitor was removed and cells were incubated with un-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Journal: Respiratory Research

    Article Title: CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts

    doi: 10.1186/s12931-021-01677-0

    Figure Lengend Snippet: Effect of ERK1/2 inhibitor (ERK1/2 inh) on growth factor expression. Primary basal cells from each of 3 non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 5 µM ERK1/2 inhibitor (LY32149966) and a combination of LPA and ERK1/2 inhibitor (LY32149966). a mRNA. Shown are expression levels of connective tissue growth factor ( CTGF ), endothelin-1 ( EDN1 ), transforming growth factor beta ( TGFB1 ) and platelet derived growth factor B ( PDGFB ) determined after 3 h of LPA exposure with and without ERK1/2 inhibitor (LY32149966) using qRT-PCR using 18S RNA to normalize the samples. b Protein. Shown are growth factor levels in the conditioned media. After 3 h of LPA with and without CREB inhibitor (666-15) exposure, LPA with or without ERK1/2 inhibitor was removed and cells were incubated with un-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Article Snippet: The human COL1A1 ELISA kit (MyBioSource, La Jolla, CA), ENPP2 human ELISA kit (R&D Systems), human LPA ELISA Kit (Echelon Biosciences) were performed according to the manufacturer’s instructions.

    Techniques: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of EGFR inhibitor (EGFR inh) on growth factor mRNA levels. Primary basal cells (passage 3) from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 10 µM EGFR inhibitor (AG1478), and a combination of LPA and EGFR inhibitor. a mRNA expression levels shown are mRNA levels of connective tissue growth factor ( CTGF ), endothelin-1( EDN1 ), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor B were determined after 3 h of LPA exposure with and without the inhibitor using qRT-PCR using 18S RNA to normalize the samples. b Protein levels in the conditioned media. After 3 h of LPA with and without EGFR inhibitor (AG1478) exposure, LPA with or without inhibitor was removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Journal: Respiratory Research

    Article Title: CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts

    doi: 10.1186/s12931-021-01677-0

    Figure Lengend Snippet: Effect of EGFR inhibitor (EGFR inh) on growth factor mRNA levels. Primary basal cells (passage 3) from each of three non-smoking donors were plated in triplicate in the presence or absence of 1 μg/ml LPA, 10 µM EGFR inhibitor (AG1478), and a combination of LPA and EGFR inhibitor. a mRNA expression levels shown are mRNA levels of connective tissue growth factor ( CTGF ), endothelin-1( EDN1 ), transforming growth factor beta ( TGFB1 ), and platelet derived growth factor B were determined after 3 h of LPA exposure with and without the inhibitor using qRT-PCR using 18S RNA to normalize the samples. b Protein levels in the conditioned media. After 3 h of LPA with and without EGFR inhibitor (AG1478) exposure, LPA with or without inhibitor was removed and cells were incubated with non-supplemented basal media for 24 h. Conditioned media was harvested and evaluated for growth factor content by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. ND—not detected. *p < 0.05, **p < 0.01, ***p < 0.001

    Article Snippet: The human COL1A1 ELISA kit (MyBioSource, La Jolla, CA), ENPP2 human ELISA kit (R&D Systems), human LPA ELISA Kit (Echelon Biosciences) were performed according to the manufacturer’s instructions.

    Techniques: Expressing, Derivative Assay, Quantitative RT-PCR, Incubation, Enzyme-linked Immunosorbent Assay

    Effect of CREB inhibitor (CREB inh) on fibroblast expression in response to basal cell conditioned medium. Primary normal human lung fibroblasts (NHLF) were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Fibroblast expression of collagen type 1 ( COL1A1 ) and smooth muscle actin ( ACTA2 ) was assessed after 24 h in conditioned medium. a mRNA levels. RNA was harvested and evaluated by qRT-PCR using 18S RNA to normalize the samples. b Protein levels. At the same time point, conditioned media was collected and assessed for COL1A1 secretion. Cells were solubilized with a RIPA buffer and evaluated for alpha smooth muscle myosin by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001

    Journal: Respiratory Research

    Article Title: CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts

    doi: 10.1186/s12931-021-01677-0

    Figure Lengend Snippet: Effect of CREB inhibitor (CREB inh) on fibroblast expression in response to basal cell conditioned medium. Primary normal human lung fibroblasts (NHLF) were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Fibroblast expression of collagen type 1 ( COL1A1 ) and smooth muscle actin ( ACTA2 ) was assessed after 24 h in conditioned medium. a mRNA levels. RNA was harvested and evaluated by qRT-PCR using 18S RNA to normalize the samples. b Protein levels. At the same time point, conditioned media was collected and assessed for COL1A1 secretion. Cells were solubilized with a RIPA buffer and evaluated for alpha smooth muscle myosin by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001

    Article Snippet: The human COL1A1 ELISA kit (MyBioSource, La Jolla, CA), ENPP2 human ELISA kit (R&D Systems), human LPA ELISA Kit (Echelon Biosciences) were performed according to the manufacturer’s instructions.

    Techniques: Expressing, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

    Effect of CREB inhibitor (CREB inh) on autotaxin ( ENPP2 ) expression, secretion, and activity in fibroblasts in response to medium conditioned by LPA-treated basal cells. Primary NHLF were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1.0 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Autotaxin, an enzyme responsible for the extracellular production of LPA, in encoded by ENPP2 . Expression of the ENPP2 gene was assessed in fibroblasts 24 h after exposure to conditioned medium. Autotaxin protein levels and LPA levels were assessed in cell culture medium 24 h after exposure to conditioned medium. a ENPP2 mRNA levels. mRNA was measured by qRT-PCR using 18S RNA to normalize the samples. b Autotaxin protein levels. Protein levels were measured in cell culture medium by ELISA. c LPA levels. LPA was measured in cell culture medium by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001

    Journal: Respiratory Research

    Article Title: CREB-dependent LPA-induced signaling initiates a pro-fibrotic feedback loop between small airway basal cells and fibroblasts

    doi: 10.1186/s12931-021-01677-0

    Figure Lengend Snippet: Effect of CREB inhibitor (CREB inh) on autotaxin ( ENPP2 ) expression, secretion, and activity in fibroblasts in response to medium conditioned by LPA-treated basal cells. Primary NHLF were treated with conditioned media from each of 3 non-smoking donors obtained in the presence or absence of 1.0 μg/ml LPA. For each condition, conditioned medium collected from cultures that were naive or treated with vehicle control (DMSO) or 200 nM CREB inhibitor (666-15) were collected and transferred to NHLF. Autotaxin, an enzyme responsible for the extracellular production of LPA, in encoded by ENPP2 . Expression of the ENPP2 gene was assessed in fibroblasts 24 h after exposure to conditioned medium. Autotaxin protein levels and LPA levels were assessed in cell culture medium 24 h after exposure to conditioned medium. a ENPP2 mRNA levels. mRNA was measured by qRT-PCR using 18S RNA to normalize the samples. b Autotaxin protein levels. Protein levels were measured in cell culture medium by ELISA. c LPA levels. LPA was measured in cell culture medium by ELISA. Data are expressed as the mean value of the 3 donors ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001

    Article Snippet: The human COL1A1 ELISA kit (MyBioSource, La Jolla, CA), ENPP2 human ELISA kit (R&D Systems), human LPA ELISA Kit (Echelon Biosciences) were performed according to the manufacturer’s instructions.

    Techniques: Expressing, Activity Assay, Cell Culture, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

    Experimental procedures used for determination of endometrial LPA, LPAR1–4 mRNA level and protein abundance, effect of LPA on PG and CTGF secretion at different stages of endometrosis. Ctr-control,  LPA-lysophosphatidic  acid, AA- arachidonic acid

    Journal: BMC Veterinary Research

    Article Title: Lysophosphatidic acid as a regulator of endometrial connective tissue growth factor and prostaglandin secretion during estrous cycle and endometrosis in the mare

    doi: 10.1186/s12917-020-02562-6

    Figure Lengend Snippet: Experimental procedures used for determination of endometrial LPA, LPAR1–4 mRNA level and protein abundance, effect of LPA on PG and CTGF secretion at different stages of endometrosis. Ctr-control, LPA-lysophosphatidic acid, AA- arachidonic acid

    Article Snippet: The concentration of LPA in endometrial tissue was determined using the Lysophosphatidic Acid Assay Kit II (Echelon Biosciences Inc.; #K-2800S) following the manufacturer’s instructions.

    Techniques:

    Lysophosphatidic acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.

    Journal: The Journal of surgical research

    Article Title: Protective Effect of Phosphatidylserine Blockade in Hemorrhagic Shock Phosphatidylserine Blockade in Hemorrhage

    doi: 10.1016/j.jss.2019.07.050

    Figure Lengend Snippet: Lysophosphatidic acid (LPA) levels in the control, hemorrhage only (HEM), and Diannexin followed by hemorrhage (DA + HEM) groups. Data were collected at the following time points: baseline, post-hemorrhage (post-hem), and post-resuscitation (post-resus). PS blockade reduced LPA levels compared to the HEM group. * denotes p<0.05 on analysis of variance among the three groups. LPA units are in μM. Error bars indicate SEM.

    Article Snippet: LPA was measured using Echelon Biosciences lysophosphatidic acid competitive ELISA kit for rat serum.

    Techniques: