peroxidase anti peroxidase igg immune complexes  (Valiant)

 
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    Name:
    Anti horseradish peroxidase rabbit IgG fraction
    Description:
    Product is the lyophilized powder of rabbit IgG fraction to horseradish peroxidase and buffer salts
    Catalog Number:
    0855974
    Price:
    267.9
    Category:
    Life Sciences Antibodies Primary Antibodies
    Applications:
    Immunoassays
    Size:
    5 mL
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    Structured Review

    Valiant peroxidase anti peroxidase igg immune complexes
    Western blot analysis for tyrosine-phosphorylated (PY) FcRγ-subunit of PMN or macrophage lysates after in vitro incubation. (A,B) 3×10 6 cells/ml from either Wt or TLR4 mut mice were incubated for 5, 15, and 30 min with <t>IgG</t> immune complexes (IgGIC; 100 µg/ml). (C,D) The same protocol was used for stimulation with LPS (20 ng/ml). (E) Lysates from either Wt or TLR4 mut mice that were incubated with polymyxin-treated <t>BSA</t> immune complexes (100 µg/ml) under the same conditions as described above. Corresponding loading controls are displayed in the lower panels.
    Product is the lyophilized powder of rabbit IgG fraction to horseradish peroxidase and buffer salts
    https://www.bioz.com/result/peroxidase anti peroxidase igg immune complexes/product/Valiant
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    peroxidase anti peroxidase igg immune complexes - by Bioz Stars, 2021-03
    93/100 stars

    Images

    1) Product Images from "Cross-Talk between TLR4 and Fc?ReceptorIII (CD16) Pathways"

    Article Title: Cross-Talk between TLR4 and Fc?ReceptorIII (CD16) Pathways

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1000464

    Western blot analysis for tyrosine-phosphorylated (PY) FcRγ-subunit of PMN or macrophage lysates after in vitro incubation. (A,B) 3×10 6 cells/ml from either Wt or TLR4 mut mice were incubated for 5, 15, and 30 min with IgG immune complexes (IgGIC; 100 µg/ml). (C,D) The same protocol was used for stimulation with LPS (20 ng/ml). (E) Lysates from either Wt or TLR4 mut mice that were incubated with polymyxin-treated BSA immune complexes (100 µg/ml) under the same conditions as described above. Corresponding loading controls are displayed in the lower panels.
    Figure Legend Snippet: Western blot analysis for tyrosine-phosphorylated (PY) FcRγ-subunit of PMN or macrophage lysates after in vitro incubation. (A,B) 3×10 6 cells/ml from either Wt or TLR4 mut mice were incubated for 5, 15, and 30 min with IgG immune complexes (IgGIC; 100 µg/ml). (C,D) The same protocol was used for stimulation with LPS (20 ng/ml). (E) Lysates from either Wt or TLR4 mut mice that were incubated with polymyxin-treated BSA immune complexes (100 µg/ml) under the same conditions as described above. Corresponding loading controls are displayed in the lower panels.

    Techniques Used: Western Blot, In Vitro, Incubation, Mouse Assay

    Parameters of acute lung injury in Wt and TLR4 mut mice. (A) Lung injury (as measured by leak of 125 I-BSA into lung) in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice receiving LPS intratracheally. (B) Permeability indices in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice after intrapulmonary immune complex formation following administration of BSA (i.v.) and anti-BSA IgG (i.t.). (C) IL-6 levels in BAL fluids after IgG immune complex (IgGIC)- or LPS-induced lung injury using Wt and TLR4 mut mice. (D) TNFα in BAL fluids from the same mice described in frame (C). For each bar, n≥5. (E) Lung injury induced by IgG immune complexes (IgGIC) in Wt and TLR4 mut mice after endotoxin removal by polymyxin. (F) Lung permeability after intratracheal (i.t.) administration of anti-BSA IgG and intravenous (i.v.) injection of BSA, PBS i.t., and BSA i.v. or anti-BSA i.t. and PBS i.v. (G) IgGIC-induced lung injury in FcRγ-subunit −/− mice in comparison to Wt mice (FcRγ-subunit +/+ ). For each bar, n≥5.
    Figure Legend Snippet: Parameters of acute lung injury in Wt and TLR4 mut mice. (A) Lung injury (as measured by leak of 125 I-BSA into lung) in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice receiving LPS intratracheally. (B) Permeability indices in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice after intrapulmonary immune complex formation following administration of BSA (i.v.) and anti-BSA IgG (i.t.). (C) IL-6 levels in BAL fluids after IgG immune complex (IgGIC)- or LPS-induced lung injury using Wt and TLR4 mut mice. (D) TNFα in BAL fluids from the same mice described in frame (C). For each bar, n≥5. (E) Lung injury induced by IgG immune complexes (IgGIC) in Wt and TLR4 mut mice after endotoxin removal by polymyxin. (F) Lung permeability after intratracheal (i.t.) administration of anti-BSA IgG and intravenous (i.v.) injection of BSA, PBS i.t., and BSA i.v. or anti-BSA i.t. and PBS i.v. (G) IgGIC-induced lung injury in FcRγ-subunit −/− mice in comparison to Wt mice (FcRγ-subunit +/+ ). For each bar, n≥5.

    Techniques Used: Mouse Assay, Permeability, Injection

    Association between TLR4 and FcRγIII. Peritoneal PMNs and macrophages (3×10 6 cells/ml) from Wt mice and FcRγ-subunit −/− mice were incubated in vitro for 30 min with either IgG immune complexes (IgGIC; 100 µg/ml), LPS (20 ng/ml), or the combination. (A,B) Western blot analysis (IB) for FcγRIII of Wt PMN or macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. (C,D) Reverse direction immunoprecipitation using anti-FcγRII/III IgG followed by Western blot analysis for TLR4. (E,F) Western blot analysis for FcγRIII of PMNs or macrophages from FcγRIII −/− co-immunoprecipitated (IP) with anti-TLR4. (G,H) Samples were immunoprecipitated with anti-TLR6 IgG and probed for FcγRIII. (I,J) Immunoprecipitation with anti-CD23 followed by Western blots using anti-TLR4 IgG. (K) Western blots (IB) of cell lysates of Wt macrophages that were incubated for 30 min with BSA IgG immune complexes (IgGIC; 100 µg/ml), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC; 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). IB for FcγRIII of Wt macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. Corresponding loading controls are displayed in lower panels.
    Figure Legend Snippet: Association between TLR4 and FcRγIII. Peritoneal PMNs and macrophages (3×10 6 cells/ml) from Wt mice and FcRγ-subunit −/− mice were incubated in vitro for 30 min with either IgG immune complexes (IgGIC; 100 µg/ml), LPS (20 ng/ml), or the combination. (A,B) Western blot analysis (IB) for FcγRIII of Wt PMN or macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. (C,D) Reverse direction immunoprecipitation using anti-FcγRII/III IgG followed by Western blot analysis for TLR4. (E,F) Western blot analysis for FcγRIII of PMNs or macrophages from FcγRIII −/− co-immunoprecipitated (IP) with anti-TLR4. (G,H) Samples were immunoprecipitated with anti-TLR6 IgG and probed for FcγRIII. (I,J) Immunoprecipitation with anti-CD23 followed by Western blots using anti-TLR4 IgG. (K) Western blots (IB) of cell lysates of Wt macrophages that were incubated for 30 min with BSA IgG immune complexes (IgGIC; 100 µg/ml), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC; 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). IB for FcγRIII of Wt macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. Corresponding loading controls are displayed in lower panels.

    Techniques Used: Mouse Assay, Incubation, In Vitro, Western Blot, Immunoprecipitation

    In vitro cytokine responses of elicited peritoneal PMNs and macrophages to LPS and IgGIC. In vitro cytokine responses of elicited peritoneal PMNs (A–D) and macrophages (E,F). Cells (3×10 6 cells/ml) from either Wt or TLR4 mut mice were incubated for 4 hr with LPS (20 ng/ml) or IgGIC; 100 µg/ml), respectively. In addition, macrophages were incubated with polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). (A) IL-6 release from PMNs after LPS stimulation. (B) TNFα levels after incubation of PMNs with LPS. (C) Concentration of IL-6 in supernatants when PMNs were exposed to IgGIC. (D) Production of TNFα by PMNs and macrophages in the presence of IgGIC. Ctrl = control levels of non-stimulated cells. (E) Release of IL-6 by macrophages into supernatant fluids after stimulation with LPS, IgGIC, p.-t. BSA IC, or PAP IC. (F) TNFα production by macrophages exposed to LPS, IgGIC, p.-t. BSA IC, or PAP IC. The experiments were performed in triplicates for each condition (each bar) with n≥3 donors of cells for each mouse strain, Wt or TLR4 mut. Differences between controls and stimulated cells were—if not otherwise noted—statistically significant (p
    Figure Legend Snippet: In vitro cytokine responses of elicited peritoneal PMNs and macrophages to LPS and IgGIC. In vitro cytokine responses of elicited peritoneal PMNs (A–D) and macrophages (E,F). Cells (3×10 6 cells/ml) from either Wt or TLR4 mut mice were incubated for 4 hr with LPS (20 ng/ml) or IgGIC; 100 µg/ml), respectively. In addition, macrophages were incubated with polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). (A) IL-6 release from PMNs after LPS stimulation. (B) TNFα levels after incubation of PMNs with LPS. (C) Concentration of IL-6 in supernatants when PMNs were exposed to IgGIC. (D) Production of TNFα by PMNs and macrophages in the presence of IgGIC. Ctrl = control levels of non-stimulated cells. (E) Release of IL-6 by macrophages into supernatant fluids after stimulation with LPS, IgGIC, p.-t. BSA IC, or PAP IC. (F) TNFα production by macrophages exposed to LPS, IgGIC, p.-t. BSA IC, or PAP IC. The experiments were performed in triplicates for each condition (each bar) with n≥3 donors of cells for each mouse strain, Wt or TLR4 mut. Differences between controls and stimulated cells were—if not otherwise noted—statistically significant (p

    Techniques Used: In Vitro, Mouse Assay, Incubation, Concentration Assay

    Related Articles

    Incubation:

    Article Title: Overexpression of Receptor for Advanced Glycation End Products and High-Mobility Group Box 1 in Human Dental Pulp Inflammation
    Article Snippet: Rabbit anti-HMGB-1 or RAGE primary antibodies were incubated for 3 h at RT and overnight at 4°C, respectively. .. Then the membranes were washed and incubated with horseradish peroxidase-conjugated anti-rabbit polyclonal IgG (MP Biomedicals Inc., Solon, OH, USA) at RT for 1 h. Labeled bands were visualized using an enhanced chemiluminescence system (GE Healthcare Bio-Science, Pittsburgh, PA, USA) and exposed to high-performance chemiluminescence film (GE Healthcare). .. The intensity of the protein bands in western blotting was quantified using National Institutes of Health Image 1.63 software.

    Article Title: Cross-Talk between TLR4 and Fc?ReceptorIII (CD16) Pathways
    Article Snippet: In Vitro Incubation of Peritoneal PMNs and Macrophages Mouse peritoneal leukocytes were harvested 5 h (PMNs) or 5 days (macrophages) after intraperitoneal injection of thioglycolate into untreated Wt and TLR4 mut mice by peritoneal lavage with PBS. .. 3×106 cells / sample were incubated in HBSS for up to 4 h at 37°C in the presence of LPS (20 ng/ml; serotype O111:B4; Sigma, St. Louis, MO), BSA IgG immune complexes (IgGIC, 100 µg/ml; MP Biomedicals), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml), peroxidase/anti-peroxidase IgG immune complexes (PAP IC, 100 µg/ml; MP Biomedicals), opsonized zymosan particles (300 µg/ml; Sigma) or Pam3Cys (1 µg/ml; InvivoGen). .. After incubation, supernatant fluids were collected for assessment of cytokines by ELISA and pellets were lysed with RIPA buffer (Upstate) for immunoprecipitation analyses.

    Labeling:

    Article Title: Overexpression of Receptor for Advanced Glycation End Products and High-Mobility Group Box 1 in Human Dental Pulp Inflammation
    Article Snippet: Rabbit anti-HMGB-1 or RAGE primary antibodies were incubated for 3 h at RT and overnight at 4°C, respectively. .. Then the membranes were washed and incubated with horseradish peroxidase-conjugated anti-rabbit polyclonal IgG (MP Biomedicals Inc., Solon, OH, USA) at RT for 1 h. Labeled bands were visualized using an enhanced chemiluminescence system (GE Healthcare Bio-Science, Pittsburgh, PA, USA) and exposed to high-performance chemiluminescence film (GE Healthcare). .. The intensity of the protein bands in western blotting was quantified using National Institutes of Health Image 1.63 software.

    other:

    Article Title: A New Highly Selective and Specific Anti-puerarin polyclonal Antibody for Determination of Puerarin Using a Mannich Reaction Hapten Conjugate
    Article Snippet: Peroxidase-labeled anti-rabbit IgG was obtained from MP Biomedicals (Ohio, USA).

    Staining:

    Article Title: Protein Interactomes Identify Distinct Pathways for Streptococcus mutans YidC1 and YidC2 Membrane Protein Insertases
    Article Snippet: Cell lysate samples were electrophoresed on 4-20% precast gels (Bio-Rad Laboratories, Hercules, CA) in Tris-Glycine-SDS buffer. .. Replicate gels were stained with Coomassie Blue R 250 or transblotted onto Immobilon polyvinylidene difluoride (PVDF) membranes (Bio-Rad Laboratories, Hercules, CA), reacted with affinity-purified YidC1 or YidC2 C-terminal-specific polyclonal rabbit antibodies (1:1000) [ ], or anti-Ffh or anti-FtsY polyclonal rabbit antisera (1:1000) [ ], followed by horseradish peroxidase-labeled anti-rabbit IgG (MP Biomedicals, Irvine, CA) (1:5000), and developed using the enhanced-chemiluminescence (ECL) Western blotting system (GE Healthcare). .. Coupling of anti-YidC2 antibodies to DynaBeads™ and immunocapture of protein complexes Five mg of M-280 Tosylactivated Dynabeads™ (Invitrogen) were washed twice with 1 ml 0.1 M Na-phosphate buffer pH 7.4.

    Affinity Purification:

    Article Title: Protein Interactomes Identify Distinct Pathways for Streptococcus mutans YidC1 and YidC2 Membrane Protein Insertases
    Article Snippet: Cell lysate samples were electrophoresed on 4-20% precast gels (Bio-Rad Laboratories, Hercules, CA) in Tris-Glycine-SDS buffer. .. Replicate gels were stained with Coomassie Blue R 250 or transblotted onto Immobilon polyvinylidene difluoride (PVDF) membranes (Bio-Rad Laboratories, Hercules, CA), reacted with affinity-purified YidC1 or YidC2 C-terminal-specific polyclonal rabbit antibodies (1:1000) [ ], or anti-Ffh or anti-FtsY polyclonal rabbit antisera (1:1000) [ ], followed by horseradish peroxidase-labeled anti-rabbit IgG (MP Biomedicals, Irvine, CA) (1:5000), and developed using the enhanced-chemiluminescence (ECL) Western blotting system (GE Healthcare). .. Coupling of anti-YidC2 antibodies to DynaBeads™ and immunocapture of protein complexes Five mg of M-280 Tosylactivated Dynabeads™ (Invitrogen) were washed twice with 1 ml 0.1 M Na-phosphate buffer pH 7.4.

    Western Blot:

    Article Title: Protein Interactomes Identify Distinct Pathways for Streptococcus mutans YidC1 and YidC2 Membrane Protein Insertases
    Article Snippet: Cell lysate samples were electrophoresed on 4-20% precast gels (Bio-Rad Laboratories, Hercules, CA) in Tris-Glycine-SDS buffer. .. Replicate gels were stained with Coomassie Blue R 250 or transblotted onto Immobilon polyvinylidene difluoride (PVDF) membranes (Bio-Rad Laboratories, Hercules, CA), reacted with affinity-purified YidC1 or YidC2 C-terminal-specific polyclonal rabbit antibodies (1:1000) [ ], or anti-Ffh or anti-FtsY polyclonal rabbit antisera (1:1000) [ ], followed by horseradish peroxidase-labeled anti-rabbit IgG (MP Biomedicals, Irvine, CA) (1:5000), and developed using the enhanced-chemiluminescence (ECL) Western blotting system (GE Healthcare). .. Coupling of anti-YidC2 antibodies to DynaBeads™ and immunocapture of protein complexes Five mg of M-280 Tosylactivated Dynabeads™ (Invitrogen) were washed twice with 1 ml 0.1 M Na-phosphate buffer pH 7.4.

    Immunohistochemistry:

    Article Title: Drosophila Embryos as Model to Assess Cellular and Developmental Toxicity of Multi-Walled Carbon Nanotubes (MWCNT) in Living Organisms
    Article Snippet: Embryos injected as control with water, 10% DMSO, unlabelled MWCNTs or with Camptothecin were treated the same way. .. Immunohistochemistry The following antibodies were used BP102 (mouse, 1∶1000; kindly provided by Professor Nipam Patel), anti- γH2Av (1∶1000; kindly provided by Kim McKim) and anti-HRP coupled to FITC (goat, 1∶500, MPbio). .. All antibodies are used for routine stains in Drosophila embryonic research and their staining pattern has been well described

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    Valiant peroxidase anti peroxidase igg immune complexes
    Western blot analysis for tyrosine-phosphorylated (PY) FcRγ-subunit of PMN or macrophage lysates after in vitro incubation. (A,B) 3×10 6 cells/ml from either Wt or TLR4 mut mice were incubated for 5, 15, and 30 min with <t>IgG</t> immune complexes (IgGIC; 100 µg/ml). (C,D) The same protocol was used for stimulation with LPS (20 ng/ml). (E) Lysates from either Wt or TLR4 mut mice that were incubated with polymyxin-treated <t>BSA</t> immune complexes (100 µg/ml) under the same conditions as described above. Corresponding loading controls are displayed in the lower panels.
    Peroxidase Anti Peroxidase Igg Immune Complexes, supplied by Valiant, 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/peroxidase anti peroxidase igg immune complexes/product/Valiant
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    peroxidase anti peroxidase igg immune complexes - by Bioz Stars, 2021-03
    93/100 stars
      Buy from Supplier

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    Western blot analysis for tyrosine-phosphorylated (PY) FcRγ-subunit of PMN or macrophage lysates after in vitro incubation. (A,B) 3×10 6 cells/ml from either Wt or TLR4 mut mice were incubated for 5, 15, and 30 min with IgG immune complexes (IgGIC; 100 µg/ml). (C,D) The same protocol was used for stimulation with LPS (20 ng/ml). (E) Lysates from either Wt or TLR4 mut mice that were incubated with polymyxin-treated BSA immune complexes (100 µg/ml) under the same conditions as described above. Corresponding loading controls are displayed in the lower panels.

    Journal: PLoS Pathogens

    Article Title: Cross-Talk between TLR4 and Fc?ReceptorIII (CD16) Pathways

    doi: 10.1371/journal.ppat.1000464

    Figure Lengend Snippet: Western blot analysis for tyrosine-phosphorylated (PY) FcRγ-subunit of PMN or macrophage lysates after in vitro incubation. (A,B) 3×10 6 cells/ml from either Wt or TLR4 mut mice were incubated for 5, 15, and 30 min with IgG immune complexes (IgGIC; 100 µg/ml). (C,D) The same protocol was used for stimulation with LPS (20 ng/ml). (E) Lysates from either Wt or TLR4 mut mice that were incubated with polymyxin-treated BSA immune complexes (100 µg/ml) under the same conditions as described above. Corresponding loading controls are displayed in the lower panels.

    Article Snippet: 3×106 cells / sample were incubated in HBSS for up to 4 h at 37°C in the presence of LPS (20 ng/ml; serotype O111:B4; Sigma, St. Louis, MO), BSA IgG immune complexes (IgGIC, 100 µg/ml; MP Biomedicals), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml), peroxidase/anti-peroxidase IgG immune complexes (PAP IC, 100 µg/ml; MP Biomedicals), opsonized zymosan particles (300 µg/ml; Sigma) or Pam3Cys (1 µg/ml; InvivoGen).

    Techniques: Western Blot, In Vitro, Incubation, Mouse Assay

    Parameters of acute lung injury in Wt and TLR4 mut mice. (A) Lung injury (as measured by leak of 125 I-BSA into lung) in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice receiving LPS intratracheally. (B) Permeability indices in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice after intrapulmonary immune complex formation following administration of BSA (i.v.) and anti-BSA IgG (i.t.). (C) IL-6 levels in BAL fluids after IgG immune complex (IgGIC)- or LPS-induced lung injury using Wt and TLR4 mut mice. (D) TNFα in BAL fluids from the same mice described in frame (C). For each bar, n≥5. (E) Lung injury induced by IgG immune complexes (IgGIC) in Wt and TLR4 mut mice after endotoxin removal by polymyxin. (F) Lung permeability after intratracheal (i.t.) administration of anti-BSA IgG and intravenous (i.v.) injection of BSA, PBS i.t., and BSA i.v. or anti-BSA i.t. and PBS i.v. (G) IgGIC-induced lung injury in FcRγ-subunit −/− mice in comparison to Wt mice (FcRγ-subunit +/+ ). For each bar, n≥5.

    Journal: PLoS Pathogens

    Article Title: Cross-Talk between TLR4 and Fc?ReceptorIII (CD16) Pathways

    doi: 10.1371/journal.ppat.1000464

    Figure Lengend Snippet: Parameters of acute lung injury in Wt and TLR4 mut mice. (A) Lung injury (as measured by leak of 125 I-BSA into lung) in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice receiving LPS intratracheally. (B) Permeability indices in Wt, TLR4 mut, TLR4 +/+ , and TLR4 −/− mice after intrapulmonary immune complex formation following administration of BSA (i.v.) and anti-BSA IgG (i.t.). (C) IL-6 levels in BAL fluids after IgG immune complex (IgGIC)- or LPS-induced lung injury using Wt and TLR4 mut mice. (D) TNFα in BAL fluids from the same mice described in frame (C). For each bar, n≥5. (E) Lung injury induced by IgG immune complexes (IgGIC) in Wt and TLR4 mut mice after endotoxin removal by polymyxin. (F) Lung permeability after intratracheal (i.t.) administration of anti-BSA IgG and intravenous (i.v.) injection of BSA, PBS i.t., and BSA i.v. or anti-BSA i.t. and PBS i.v. (G) IgGIC-induced lung injury in FcRγ-subunit −/− mice in comparison to Wt mice (FcRγ-subunit +/+ ). For each bar, n≥5.

    Article Snippet: 3×106 cells / sample were incubated in HBSS for up to 4 h at 37°C in the presence of LPS (20 ng/ml; serotype O111:B4; Sigma, St. Louis, MO), BSA IgG immune complexes (IgGIC, 100 µg/ml; MP Biomedicals), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml), peroxidase/anti-peroxidase IgG immune complexes (PAP IC, 100 µg/ml; MP Biomedicals), opsonized zymosan particles (300 µg/ml; Sigma) or Pam3Cys (1 µg/ml; InvivoGen).

    Techniques: Mouse Assay, Permeability, Injection

    Association between TLR4 and FcRγIII. Peritoneal PMNs and macrophages (3×10 6 cells/ml) from Wt mice and FcRγ-subunit −/− mice were incubated in vitro for 30 min with either IgG immune complexes (IgGIC; 100 µg/ml), LPS (20 ng/ml), or the combination. (A,B) Western blot analysis (IB) for FcγRIII of Wt PMN or macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. (C,D) Reverse direction immunoprecipitation using anti-FcγRII/III IgG followed by Western blot analysis for TLR4. (E,F) Western blot analysis for FcγRIII of PMNs or macrophages from FcγRIII −/− co-immunoprecipitated (IP) with anti-TLR4. (G,H) Samples were immunoprecipitated with anti-TLR6 IgG and probed for FcγRIII. (I,J) Immunoprecipitation with anti-CD23 followed by Western blots using anti-TLR4 IgG. (K) Western blots (IB) of cell lysates of Wt macrophages that were incubated for 30 min with BSA IgG immune complexes (IgGIC; 100 µg/ml), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC; 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). IB for FcγRIII of Wt macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. Corresponding loading controls are displayed in lower panels.

    Journal: PLoS Pathogens

    Article Title: Cross-Talk between TLR4 and Fc?ReceptorIII (CD16) Pathways

    doi: 10.1371/journal.ppat.1000464

    Figure Lengend Snippet: Association between TLR4 and FcRγIII. Peritoneal PMNs and macrophages (3×10 6 cells/ml) from Wt mice and FcRγ-subunit −/− mice were incubated in vitro for 30 min with either IgG immune complexes (IgGIC; 100 µg/ml), LPS (20 ng/ml), or the combination. (A,B) Western blot analysis (IB) for FcγRIII of Wt PMN or macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. (C,D) Reverse direction immunoprecipitation using anti-FcγRII/III IgG followed by Western blot analysis for TLR4. (E,F) Western blot analysis for FcγRIII of PMNs or macrophages from FcγRIII −/− co-immunoprecipitated (IP) with anti-TLR4. (G,H) Samples were immunoprecipitated with anti-TLR6 IgG and probed for FcγRIII. (I,J) Immunoprecipitation with anti-CD23 followed by Western blots using anti-TLR4 IgG. (K) Western blots (IB) of cell lysates of Wt macrophages that were incubated for 30 min with BSA IgG immune complexes (IgGIC; 100 µg/ml), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC; 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). IB for FcγRIII of Wt macrophage lysates co-immunoprecipitated (IP) with anti-TLR4. Corresponding loading controls are displayed in lower panels.

    Article Snippet: 3×106 cells / sample were incubated in HBSS for up to 4 h at 37°C in the presence of LPS (20 ng/ml; serotype O111:B4; Sigma, St. Louis, MO), BSA IgG immune complexes (IgGIC, 100 µg/ml; MP Biomedicals), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml), peroxidase/anti-peroxidase IgG immune complexes (PAP IC, 100 µg/ml; MP Biomedicals), opsonized zymosan particles (300 µg/ml; Sigma) or Pam3Cys (1 µg/ml; InvivoGen).

    Techniques: Mouse Assay, Incubation, In Vitro, Western Blot, Immunoprecipitation

    In vitro cytokine responses of elicited peritoneal PMNs and macrophages to LPS and IgGIC. In vitro cytokine responses of elicited peritoneal PMNs (A–D) and macrophages (E,F). Cells (3×10 6 cells/ml) from either Wt or TLR4 mut mice were incubated for 4 hr with LPS (20 ng/ml) or IgGIC; 100 µg/ml), respectively. In addition, macrophages were incubated with polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). (A) IL-6 release from PMNs after LPS stimulation. (B) TNFα levels after incubation of PMNs with LPS. (C) Concentration of IL-6 in supernatants when PMNs were exposed to IgGIC. (D) Production of TNFα by PMNs and macrophages in the presence of IgGIC. Ctrl = control levels of non-stimulated cells. (E) Release of IL-6 by macrophages into supernatant fluids after stimulation with LPS, IgGIC, p.-t. BSA IC, or PAP IC. (F) TNFα production by macrophages exposed to LPS, IgGIC, p.-t. BSA IC, or PAP IC. The experiments were performed in triplicates for each condition (each bar) with n≥3 donors of cells for each mouse strain, Wt or TLR4 mut. Differences between controls and stimulated cells were—if not otherwise noted—statistically significant (p

    Journal: PLoS Pathogens

    Article Title: Cross-Talk between TLR4 and Fc?ReceptorIII (CD16) Pathways

    doi: 10.1371/journal.ppat.1000464

    Figure Lengend Snippet: In vitro cytokine responses of elicited peritoneal PMNs and macrophages to LPS and IgGIC. In vitro cytokine responses of elicited peritoneal PMNs (A–D) and macrophages (E,F). Cells (3×10 6 cells/ml) from either Wt or TLR4 mut mice were incubated for 4 hr with LPS (20 ng/ml) or IgGIC; 100 µg/ml), respectively. In addition, macrophages were incubated with polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml) or peroxidase/anti-peroxidase IgGIC immune complexes (PAP IC, 100 µg/ml). (A) IL-6 release from PMNs after LPS stimulation. (B) TNFα levels after incubation of PMNs with LPS. (C) Concentration of IL-6 in supernatants when PMNs were exposed to IgGIC. (D) Production of TNFα by PMNs and macrophages in the presence of IgGIC. Ctrl = control levels of non-stimulated cells. (E) Release of IL-6 by macrophages into supernatant fluids after stimulation with LPS, IgGIC, p.-t. BSA IC, or PAP IC. (F) TNFα production by macrophages exposed to LPS, IgGIC, p.-t. BSA IC, or PAP IC. The experiments were performed in triplicates for each condition (each bar) with n≥3 donors of cells for each mouse strain, Wt or TLR4 mut. Differences between controls and stimulated cells were—if not otherwise noted—statistically significant (p

    Article Snippet: 3×106 cells / sample were incubated in HBSS for up to 4 h at 37°C in the presence of LPS (20 ng/ml; serotype O111:B4; Sigma, St. Louis, MO), BSA IgG immune complexes (IgGIC, 100 µg/ml; MP Biomedicals), polymyxin-treated BSA IgG immune complexes (p.-t. BSA IC, 100 µg/ml), peroxidase/anti-peroxidase IgG immune complexes (PAP IC, 100 µg/ml; MP Biomedicals), opsonized zymosan particles (300 µg/ml; Sigma) or Pam3Cys (1 µg/ml; InvivoGen).

    Techniques: In Vitro, Mouse Assay, Incubation, Concentration Assay