hek 293 cells mycoplasma free Search Results


  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 86
    GenScript corporation eukaryotic mycoplasma free hek 293 cells
    Gliadin-induced innate immune responses are elicited by wheat ATI, a protein copurifying with ω-gliadins. (A) Stimulation of THP-1 cells with α-, γ-, ω1.2-, and ω5-gliadin fractions (all 100 µg/ml) isolated from the pure wheat strain Rektor. Co-incubation of α- and γ-gliadin with 100 µg/ml of regular PT gliadin from Sigma-Aldrich served as cell viability control. LPS was used as positive control, whereas PT or PT zein served as negative control. (B and C) IL-8 secretion after stimulation with 100 µg/ml ω-gliadins in TLR4-transfected (B) and in untransfected <t>HEK-293</t> cells (C). 10 ng/ml PMA served as cell viability control. 10 ng/ml LPS, 100 µg/ml PT gliadin, or 100 µg/ml of a PT digest of Rektor gliadin (PT Rektor) served as positive control, and 100 µg/ml PT zein, 1 µg/ml Pam3CSK4, or a PT mixture (PT ctrl) served as negative controls. (D) Stimulatory capacity of synthetic overlapping 20mers of ω5-gliadin in TLR4-transfected HEK-293 cells. For illustration purposes, 9 fractions each were pooled in the stimulation experiments (each fraction at a concentration of 100 µg/ml), while also all 43 fractions were tested individually. LPS served as positive and Pam3CSK4 or PT zein as negative controls. (E and F) Dose response of IL-8 release by monocyte-derived DCs stimulated with water-soluble (ws) gliadin (which is enriched in ATI; E) or with purified ATI (F). LPS and water-soluble zein served as positive and negative controls, respectively. (G) Secretion of IL-12 in monocyte-derived DCs from healthy subjects upon stimulation with ATI and PT gliadin in the presence of 1,000 U/ml Interferon-γ as co-stimulatory protein. LPS and PT zein served as positive and negative controls, respectively. (H) Effect of proteinase K digestion of ATI and LPS on IL-8 secretion in DCs. (I) KC secretion in peritoneal macrophages isolated from MyD88 −/− mice compared with C57BL/6J wild-type mice upon ATI or water-soluble gliadin stimulation. LPS and water-soluble zein served as positive and negative controls, respectively. (J) IL-8 secretion of monocyte-derived DCs stimulated with ATI and LPS after preincubation with anti-TLR4 or anti-CD14 antibodies. TLR2 agonist Pam3CSK4 served as positive control. *, P < 0.05 versus negative (positive) control (if not indicated otherwise); all graphs illustrate representative data from one of at least three independent experiments, all performed in triplicates. Error bars depict standard errors of the mean.
    Eukaryotic Mycoplasma Free Hek 293 Cells, supplied by GenScript corporation, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/eukaryotic mycoplasma free hek 293 cells/product/GenScript corporation
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    eukaryotic mycoplasma free hek 293 cells - by Bioz Stars, 2024-07
    86/100 stars
      Buy from Supplier

    Image Search Results


    Gliadin-induced innate immune responses are elicited by wheat ATI, a protein copurifying with ω-gliadins. (A) Stimulation of THP-1 cells with α-, γ-, ω1.2-, and ω5-gliadin fractions (all 100 µg/ml) isolated from the pure wheat strain Rektor. Co-incubation of α- and γ-gliadin with 100 µg/ml of regular PT gliadin from Sigma-Aldrich served as cell viability control. LPS was used as positive control, whereas PT or PT zein served as negative control. (B and C) IL-8 secretion after stimulation with 100 µg/ml ω-gliadins in TLR4-transfected (B) and in untransfected HEK-293 cells (C). 10 ng/ml PMA served as cell viability control. 10 ng/ml LPS, 100 µg/ml PT gliadin, or 100 µg/ml of a PT digest of Rektor gliadin (PT Rektor) served as positive control, and 100 µg/ml PT zein, 1 µg/ml Pam3CSK4, or a PT mixture (PT ctrl) served as negative controls. (D) Stimulatory capacity of synthetic overlapping 20mers of ω5-gliadin in TLR4-transfected HEK-293 cells. For illustration purposes, 9 fractions each were pooled in the stimulation experiments (each fraction at a concentration of 100 µg/ml), while also all 43 fractions were tested individually. LPS served as positive and Pam3CSK4 or PT zein as negative controls. (E and F) Dose response of IL-8 release by monocyte-derived DCs stimulated with water-soluble (ws) gliadin (which is enriched in ATI; E) or with purified ATI (F). LPS and water-soluble zein served as positive and negative controls, respectively. (G) Secretion of IL-12 in monocyte-derived DCs from healthy subjects upon stimulation with ATI and PT gliadin in the presence of 1,000 U/ml Interferon-γ as co-stimulatory protein. LPS and PT zein served as positive and negative controls, respectively. (H) Effect of proteinase K digestion of ATI and LPS on IL-8 secretion in DCs. (I) KC secretion in peritoneal macrophages isolated from MyD88 −/− mice compared with C57BL/6J wild-type mice upon ATI or water-soluble gliadin stimulation. LPS and water-soluble zein served as positive and negative controls, respectively. (J) IL-8 secretion of monocyte-derived DCs stimulated with ATI and LPS after preincubation with anti-TLR4 or anti-CD14 antibodies. TLR2 agonist Pam3CSK4 served as positive control. *, P < 0.05 versus negative (positive) control (if not indicated otherwise); all graphs illustrate representative data from one of at least three independent experiments, all performed in triplicates. Error bars depict standard errors of the mean.

    Journal: The Journal of Experimental Medicine

    Article Title: Wheat amylase trypsin inhibitors drive intestinal inflammation via activation of toll-like receptor 4

    doi: 10.1084/jem.20102660

    Figure Lengend Snippet: Gliadin-induced innate immune responses are elicited by wheat ATI, a protein copurifying with ω-gliadins. (A) Stimulation of THP-1 cells with α-, γ-, ω1.2-, and ω5-gliadin fractions (all 100 µg/ml) isolated from the pure wheat strain Rektor. Co-incubation of α- and γ-gliadin with 100 µg/ml of regular PT gliadin from Sigma-Aldrich served as cell viability control. LPS was used as positive control, whereas PT or PT zein served as negative control. (B and C) IL-8 secretion after stimulation with 100 µg/ml ω-gliadins in TLR4-transfected (B) and in untransfected HEK-293 cells (C). 10 ng/ml PMA served as cell viability control. 10 ng/ml LPS, 100 µg/ml PT gliadin, or 100 µg/ml of a PT digest of Rektor gliadin (PT Rektor) served as positive control, and 100 µg/ml PT zein, 1 µg/ml Pam3CSK4, or a PT mixture (PT ctrl) served as negative controls. (D) Stimulatory capacity of synthetic overlapping 20mers of ω5-gliadin in TLR4-transfected HEK-293 cells. For illustration purposes, 9 fractions each were pooled in the stimulation experiments (each fraction at a concentration of 100 µg/ml), while also all 43 fractions were tested individually. LPS served as positive and Pam3CSK4 or PT zein as negative controls. (E and F) Dose response of IL-8 release by monocyte-derived DCs stimulated with water-soluble (ws) gliadin (which is enriched in ATI; E) or with purified ATI (F). LPS and water-soluble zein served as positive and negative controls, respectively. (G) Secretion of IL-12 in monocyte-derived DCs from healthy subjects upon stimulation with ATI and PT gliadin in the presence of 1,000 U/ml Interferon-γ as co-stimulatory protein. LPS and PT zein served as positive and negative controls, respectively. (H) Effect of proteinase K digestion of ATI and LPS on IL-8 secretion in DCs. (I) KC secretion in peritoneal macrophages isolated from MyD88 −/− mice compared with C57BL/6J wild-type mice upon ATI or water-soluble gliadin stimulation. LPS and water-soluble zein served as positive and negative controls, respectively. (J) IL-8 secretion of monocyte-derived DCs stimulated with ATI and LPS after preincubation with anti-TLR4 or anti-CD14 antibodies. TLR2 agonist Pam3CSK4 served as positive control. *, P < 0.05 versus negative (positive) control (if not indicated otherwise); all graphs illustrate representative data from one of at least three independent experiments, all performed in triplicates. Error bars depict standard errors of the mean.

    Article Snippet: To exclude bacterial contaminants, recombinant flag-tagged ATI CM3 and 0.19 were generated in eukaryotic mycoplasma-free HEK-293 cells, using cDNAs optimized to fit eukaryotic codon usage (Genscript; CM3 and 0.19 with GenBank accession numbers AY436554.1 and AY729672.1 , respectively).

    Techniques: Isolation, Incubation, Positive Control, Negative Control, Transfection, Concentration Assay, Derivative Assay, Purification

    Identification of ATI CM3 and 0.19 as inducers of innate immune responses. (A) Coimmunoprecipitation of a soluble flag-tagged TLR4/MD2 fusion protein with or without biotinylated ATI. Pull-down was performed by streptavidin-agarose. Western blot (WB) analysis using a rabbit anti-flag antibody was performed to detect the TLR4/MD2 fusion protein. The image illustrates representative data from one of three independent experiments. (B) CM3 and 0.19 were expressed in HEK-293 cells and purified as Flag-tagged molecules, yielding expected molecular masses of 15 kD and 17 kD, respectively, after 15% SDS-PAGE and Western blotting with Flag antibody. There was no expression in mock (vector only)-transfected cells. 0.19 was not secreted into media but could be recovered in SDS/DTT buffer (Cell), deoxycholate and Triton X-100 buffer (Sup), or the remaining cell pellet. (C) Purity of CM3 and 0.19 ATI after affinity purification on Flag-agarose (15% SDS-PAGE) after staining with Coomassie blue or blotting with Flag antibody. (B and C) Arrows denote expected molecular masses of CM3 and 0.19 ATIs. (D) IL-8 secretion by TLR4–CD14–MD2-expressing HEK-293 cells after stimulation with recombinant ATI. Cells were left untreated or stimulated with affinity-purified detergent extracts from mock-transfected cells, from recombinant CM3, 0.19, or with ATI isolated from wheat (all 5 µg/ml). Controls were 5 µg/ml Flag peptide, 100 µg/ml PT gliadin, and 10 ng/ml LPS. (E) Effect of expression of CM3 and 0.19 in HEK-TLR4 cells on downstream signaling of the canonical (pNF-κB/p65) and the alternative (pIRF3) pathways. Western blots of whole cell lysates (50 µg protein). (F) Reduction and alkylation (R/A) of enriched ATIs from spring wheat (ABic: ammonium bicarbonate extract; AA: acetic acid extract). Stimulatory capacity was measured by IL-8 secretion in U-937 cells. LPS served as positive control. *, P < 0.05 versus negative (positive) control. All graphs illustrate representative data from one of at least three independent experiments. D and F were performed in triplicates. Error bars depict standard errors of the mean.

    Journal: The Journal of Experimental Medicine

    Article Title: Wheat amylase trypsin inhibitors drive intestinal inflammation via activation of toll-like receptor 4

    doi: 10.1084/jem.20102660

    Figure Lengend Snippet: Identification of ATI CM3 and 0.19 as inducers of innate immune responses. (A) Coimmunoprecipitation of a soluble flag-tagged TLR4/MD2 fusion protein with or without biotinylated ATI. Pull-down was performed by streptavidin-agarose. Western blot (WB) analysis using a rabbit anti-flag antibody was performed to detect the TLR4/MD2 fusion protein. The image illustrates representative data from one of three independent experiments. (B) CM3 and 0.19 were expressed in HEK-293 cells and purified as Flag-tagged molecules, yielding expected molecular masses of 15 kD and 17 kD, respectively, after 15% SDS-PAGE and Western blotting with Flag antibody. There was no expression in mock (vector only)-transfected cells. 0.19 was not secreted into media but could be recovered in SDS/DTT buffer (Cell), deoxycholate and Triton X-100 buffer (Sup), or the remaining cell pellet. (C) Purity of CM3 and 0.19 ATI after affinity purification on Flag-agarose (15% SDS-PAGE) after staining with Coomassie blue or blotting with Flag antibody. (B and C) Arrows denote expected molecular masses of CM3 and 0.19 ATIs. (D) IL-8 secretion by TLR4–CD14–MD2-expressing HEK-293 cells after stimulation with recombinant ATI. Cells were left untreated or stimulated with affinity-purified detergent extracts from mock-transfected cells, from recombinant CM3, 0.19, or with ATI isolated from wheat (all 5 µg/ml). Controls were 5 µg/ml Flag peptide, 100 µg/ml PT gliadin, and 10 ng/ml LPS. (E) Effect of expression of CM3 and 0.19 in HEK-TLR4 cells on downstream signaling of the canonical (pNF-κB/p65) and the alternative (pIRF3) pathways. Western blots of whole cell lysates (50 µg protein). (F) Reduction and alkylation (R/A) of enriched ATIs from spring wheat (ABic: ammonium bicarbonate extract; AA: acetic acid extract). Stimulatory capacity was measured by IL-8 secretion in U-937 cells. LPS served as positive control. *, P < 0.05 versus negative (positive) control. All graphs illustrate representative data from one of at least three independent experiments. D and F were performed in triplicates. Error bars depict standard errors of the mean.

    Article Snippet: To exclude bacterial contaminants, recombinant flag-tagged ATI CM3 and 0.19 were generated in eukaryotic mycoplasma-free HEK-293 cells, using cDNAs optimized to fit eukaryotic codon usage (Genscript; CM3 and 0.19 with GenBank accession numbers AY436554.1 and AY729672.1 , respectively).

    Techniques: Western Blot, Purification, SDS Page, Expressing, Plasmid Preparation, Transfection, Affinity Purification, Staining, Recombinant, Isolation, Positive Control