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  • 93
    Thermo Fisher igg2c
    Igg2c, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 151 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Becton Dickinson igg2c
    Restricting feeding to the day reverses the gut clock and impairs worm expulsion. ( A ) Schematic of the experimental design - food availability (marked F) was restricted to a 6 h window either mid-day (Day Fed) or mid-night (Night Fed). This restriction was in place 14 days prior to and 7 days into T . muris infection. Both groups of animals were infected at ZT0 and culled 21 days post infection. ( B ) After 14 days of restricted feeding a cohort of animals were sacrificed at ZT0 or ZT12 to assess rhythms in clock gene expression in peripheral tissues (gut) via QPCR, n = 6/group, Two way ANOVA and post hoc Tukey. ( C ) Worm burden was assessed in gut 21 days post infection, value presented is median n = 10–12, Mann Whitney test. Parasite specific IgG1 ( D ) and <t>IgG2c</t> ( E ) production on day 21, n = 12, data shown is dilution 1:120. Values shown are means, unpaired T test. See also Figs S2 and S3 .
    Igg2c, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 92/100, based on 152 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    SouthernBiotech igg2c hrp
    IFN-γ–mediated expression of T-bet promotes <t>IgG2c</t> CSR but is not required for spontaneous GC formation. (A) Representative histograms of intracellular T-bet staining of WT B cells stimulated with R848, <t>anti-IgM,</t> anti-CD40 with or without exogenous IFN-β or IFN-γ. The gray histogram indicates unstimulated WT B cells. (B) T-bet mean fluorescence intensity (MFI) in stimulated B cells. (A and B) Data are representative of two or more independent experiments. (C, left) Representative histogram of Was −/− chimera intranuclear T-bet expression in GC (PNA + GL7 + ) versus naive (non-GC; PNA − GL7 − ) WT B cells. The gray histogram indicates Tbx21 −/− B cells. (Right) T-bet mean fluorescence intensity in total B cells from WT ( n = 2) and Was −/− ( n = 7) chimeras, as well as naive and GC B cells from Was −/− chimeras. (D) Anti-dsDNA (left) and Sm/RNP (right) isotype and subclass-specific auto-Abs (normalized to mean Was −/− chimera titer). (E–H) Spleen weight (E) and number of splenic CD4 + T cells (F), PNA + FAS + GC B cells (G), and CXCR5 + PD1 + Tfh cells (H). (I) Representative splenic sections stained with B220, PNA, and CD3 demonstrating PNA + GCs in both Was −/− and B cell–intrinsic Tbx21 −/− .Was −/− chimeras. Bars, 100 µm. (D–H) Data are representative of nine WT ( n = 16), nine Was −/− ( n = 34), and three B cell–intrinsic Was −/− .Tbx21 −/− ( n = 11) independent chimeras sacrificed at 24 wk. Error bars indicate SEM. *, P
    Igg2c Hrp, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 96/100, based on 250 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    igg2c  (Abcam)
    93
    Abcam igg2c
    Tfr cells regulate the production of anti-SRBC IgG1 and anti-SRBC IgG2b. Control and Stat3FC mice were immunized with SRBC via i.p. injection. Serum samples were collected at 25 dpi. Anti-SRBC IgM, IgG1, IgG2b, <t>IgG2c,</t> IgG3 and IgA titers are shown. SRBC-specific IgD and IgE were not detectable in the serum. The X-axis shows the dilution factors. Graphs show mean +/- SEM, n = 4. ** p
    Igg2c, supplied by Abcam, used in various techniques. Bioz Stars score: 93/100, based on 104 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    SouthernBiotech goat anti mouse igg2c hrp
    Tfr cells regulate the production of anti-SRBC IgG1 and anti-SRBC IgG2b. Control and Stat3FC mice were immunized with SRBC via i.p. injection. Serum samples were collected at 25 dpi. Anti-SRBC IgM, IgG1, IgG2b, <t>IgG2c,</t> IgG3 and IgA titers are shown. SRBC-specific IgD and IgE were not detectable in the serum. The X-axis shows the dilution factors. Graphs show mean +/- SEM, n = 4. ** p
    Goat Anti Mouse Igg2c Hrp, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 97/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    SouthernBiotech goat anti mouse igg2c
    Tfr cells regulate the production of anti-SRBC IgG1 and anti-SRBC IgG2b. Control and Stat3FC mice were immunized with SRBC via i.p. injection. Serum samples were collected at 25 dpi. Anti-SRBC IgM, IgG1, IgG2b, <t>IgG2c,</t> IgG3 and IgA titers are shown. SRBC-specific IgD and IgE were not detectable in the serum. The X-axis shows the dilution factors. Graphs show mean +/- SEM, n = 4. ** p
    Goat Anti Mouse Igg2c, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 93/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    GeneTex igg2c
    CTLA-4 blockade amplifies serum HIV Gag- and Env-specific antibody responses. Post-prime serum was collected 13 days after the first VLP immunization; Post-boost 1 serum was collected 11–13 days after the second VLP immunization. Post-prime and Post-boost 1 time points analyzed serum from 10 mice per immunization group from Cohorts 1 and cohorts 2. The Post-boost 2 time point analyzed serum from the 15 mice (5 per group) in Cohort 2 and was collected 7 days after the third VLP immunization. ( A ) Total Env-specific IgG and ( B ) total Gag-specific IgG were analyzed through ELISA. Quantitative ELISAs with standard curves for IgG1, IgG2b, <t>IgG2c,</t> and IgG3 were performed on all serum samples from Cohorts 1 and 2 to detect HIV Env- and Gag-specific IgG subtypes. Shown at each of the following time points: ( C ) post-prime, ( D ) Env post-prime Gag, ( E ) post-boost 1 Env, ( F ) post-boost 1 Gag, ( G ) post-boost 2 Env, and ( H ) post-boost 2 Gag IgG subtypes. The results above are the cumulative results of two independent experiments. P-values were determined by one-way ANOVA and Tukey post-hoc analysis for multiple comparisons. A line on top of two groups indicates statistical differences between these two groups. **** p
    Igg2c, supplied by GeneTex, used in various techniques. Bioz Stars score: 91/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Thermo Fisher igg2c antibodies
    c1(70–100) dKI mice show a specific breach of B cell anergy to ssDNA. Serum from 8M B6 (circles), c1(96–100) (squares) and c1(70–100) (triangles) WT (filled) and dKI (open) mice was diluted 1:100 and assessed by ELISA for production of (A) anti-ssDNA and (B) anti-dsDNA IgM, IgM a/b , IgG, IgG2a/IgG2a a and <t>IgG2c/IgG2a</t> b autoAbs. Scatterplots show data from multiple independent experiments with n = 3–16 mice each. Symbols represent individual mice and lines show the median. Kruskal-Wallis non-parametric tests with Dunn’s post-test were used to compare B6 with c1 animals within each genotype. *p
    Igg2c Antibodies, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 96/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    SouthernBiotech anti mouse igg2c hrp
    c1(70–100) dKI mice show a specific breach of B cell anergy to ssDNA. Serum from 8M B6 (circles), c1(96–100) (squares) and c1(70–100) (triangles) WT (filled) and dKI (open) mice was diluted 1:100 and assessed by ELISA for production of (A) anti-ssDNA and (B) anti-dsDNA IgM, IgM a/b , IgG, IgG2a/IgG2a a and <t>IgG2c/IgG2a</t> b autoAbs. Scatterplots show data from multiple independent experiments with n = 3–16 mice each. Symbols represent individual mice and lines show the median. Kruskal-Wallis non-parametric tests with Dunn’s post-test were used to compare B6 with c1 animals within each genotype. *p
    Anti Mouse Igg2c Hrp, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 94/100, based on 28 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    SouthernBiotech igg2c antibodies
    c1(70–100) dKI mice show a specific breach of B cell anergy to ssDNA. Serum from 8M B6 (circles), c1(96–100) (squares) and c1(70–100) (triangles) WT (filled) and dKI (open) mice was diluted 1:100 and assessed by ELISA for production of (A) anti-ssDNA and (B) anti-dsDNA IgM, IgM a/b , IgG, IgG2a/IgG2a a and <t>IgG2c/IgG2a</t> b autoAbs. Scatterplots show data from multiple independent experiments with n = 3–16 mice each. Symbols represent individual mice and lines show the median. Kruskal-Wallis non-parametric tests with Dunn’s post-test were used to compare B6 with c1 animals within each genotype. *p
    Igg2c Antibodies, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 93/100, based on 42 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Corning Life Sciences igg2c
    Hemagglutinin (HA)- and neuraminidase (NA)-specific antibody responses in vivo . Levels of HA from Cal7-specific total IgG (A) , IgG1 (B) , and <t>IgG2c</t> (C) ; of NA from Cal7-specific total IgG (D) , IgG1 (E) , and IgG2c (F) ; and of HA from PR8-specific total IgG (G) , IgG1 (H) , and IgG2c (I) in plasma were evaluated by using ELISA 7 days after final immunization. The same plasma samples as used in Figure 4 were used here. We used 800- (•), 4000- (■), and 20,000- (▴) fold diluted plasma samples for (A–F) and 32- (•), 160- (■), and 800- (▴) fold diluted plasma samples for (G–I) . n = 5. Data are means ± SD. Significant differences were analyzed only in the 800-fold-diluted plasma samples (A–F) and the 32-fold-diluted plasma samples (G–I) . † P
    Igg2c, supplied by Corning Life Sciences, used in various techniques. Bioz Stars score: 92/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Bethyl goat anti mouse igg2c
    rAd-mIL-28B induced the reduction of anti-Ag85B antibodies in mice primed with BCG and boosted with subunit vaccine. Mice were immunized according to . At week 31, the mice were bled to confirm levels of the anti-Ag85B antibodies <t>IgG2c</t> and IgG1
    Goat Anti Mouse Igg2c, supplied by Bethyl, used in various techniques. Bioz Stars score: 92/100, based on 9 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Santa Cruz Biotechnology igg2c
    rAd-mIL-28B induced the reduction of anti-Ag85B antibodies in mice primed with BCG and boosted with subunit vaccine. Mice were immunized according to . At week 31, the mice were bled to confirm levels of the anti-Ag85B antibodies <t>IgG2c</t> and IgG1
    Igg2c, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 93/100, based on 77 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Becton Dickinson monoclonal igg2c
    rAd-mIL-28B induced the reduction of anti-Ag85B antibodies in mice primed with BCG and boosted with subunit vaccine. Mice were immunized according to . At week 31, the mice were bled to confirm levels of the anti-Ag85B antibodies <t>IgG2c</t> and IgG1
    Monoclonal Igg2c, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 85/100, based on 17 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Bio-Rad goat anti mouse igg2c
    rAd-mIL-28B induced the reduction of anti-Ag85B antibodies in mice primed with BCG and boosted with subunit vaccine. Mice were immunized according to . At week 31, the mice were bled to confirm levels of the anti-Ag85B antibodies <t>IgG2c</t> and IgG1
    Goat Anti Mouse Igg2c, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 93/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Becton Dickinson rat igg2c
    rAd-mIL-28B induced the reduction of anti-Ag85B antibodies in mice primed with BCG and boosted with subunit vaccine. Mice were immunized according to . At week 31, the mice were bled to confirm levels of the anti-Ag85B antibodies <t>IgG2c</t> and IgG1
    Rat Igg2c, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 93/100, based on 11 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Binding Site Inc igg2c
    Serum IgG1 and IgG2a (A) and IgG2b and <t>IgG2c</t> (B) responses to rHag B in rats immunized s.c. with rHag B (group A) and in rats immunized s.c. with rHag B and orally challenged with freshly harvested P. gingivalis 33277 (group B). Values are the geometric means ×/÷ SD of antibody activity in serum samples from six rats/group. The results are from one experiment but representative of two separate experiments.
    Igg2c, supplied by Binding Site Inc, used in various techniques. Bioz Stars score: 92/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Bio-Rad igg2c
    Effect of nasal administration of pCol (24-38) in groups of WKY rats ( n = 5 to 6) after the onset of disease on levels of circulating IgG1 ( A ), IgG2a ( B ), IgG2b ( C ), and <t>IgG2c</t> ( D ) anti-α3(IV)NC1 antibody concentrations. Results shown represent the mean ± SD of each group at day 28 after immunization.
    Igg2c, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 91/100, based on 15 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Bio-Rad hrp conjugated goat anti mouse igg2c antibody
    Effect of nasal administration of pCol (24-38) in groups of WKY rats ( n = 5 to 6) after the onset of disease on levels of circulating IgG1 ( A ), IgG2a ( B ), IgG2b ( C ), and <t>IgG2c</t> ( D ) anti-α3(IV)NC1 antibody concentrations. Results shown represent the mean ± SD of each group at day 28 after immunization.
    Hrp Conjugated Goat Anti Mouse Igg2c Antibody, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 92/100, based on 12 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    GeneTex hrp conjugated goat anti mouse igg2c
    Effect of nasal administration of pCol (24-38) in groups of WKY rats ( n = 5 to 6) after the onset of disease on levels of circulating IgG1 ( A ), IgG2a ( B ), IgG2b ( C ), and <t>IgG2c</t> ( D ) anti-α3(IV)NC1 antibody concentrations. Results shown represent the mean ± SD of each group at day 28 after immunization.
    Hrp Conjugated Goat Anti Mouse Igg2c, supplied by GeneTex, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Restricting feeding to the day reverses the gut clock and impairs worm expulsion. ( A ) Schematic of the experimental design - food availability (marked F) was restricted to a 6 h window either mid-day (Day Fed) or mid-night (Night Fed). This restriction was in place 14 days prior to and 7 days into T . muris infection. Both groups of animals were infected at ZT0 and culled 21 days post infection. ( B ) After 14 days of restricted feeding a cohort of animals were sacrificed at ZT0 or ZT12 to assess rhythms in clock gene expression in peripheral tissues (gut) via QPCR, n = 6/group, Two way ANOVA and post hoc Tukey. ( C ) Worm burden was assessed in gut 21 days post infection, value presented is median n = 10–12, Mann Whitney test. Parasite specific IgG1 ( D ) and IgG2c ( E ) production on day 21, n = 12, data shown is dilution 1:120. Values shown are means, unpaired T test. See also Figs S2 and S3 .

    Journal: Scientific Reports

    Article Title: The circadian regulator BMAL1 programmes responses to parasitic worm infection via a dendritic cell clock

    doi: 10.1038/s41598-018-22021-5

    Figure Lengend Snippet: Restricting feeding to the day reverses the gut clock and impairs worm expulsion. ( A ) Schematic of the experimental design - food availability (marked F) was restricted to a 6 h window either mid-day (Day Fed) or mid-night (Night Fed). This restriction was in place 14 days prior to and 7 days into T . muris infection. Both groups of animals were infected at ZT0 and culled 21 days post infection. ( B ) After 14 days of restricted feeding a cohort of animals were sacrificed at ZT0 or ZT12 to assess rhythms in clock gene expression in peripheral tissues (gut) via QPCR, n = 6/group, Two way ANOVA and post hoc Tukey. ( C ) Worm burden was assessed in gut 21 days post infection, value presented is median n = 10–12, Mann Whitney test. Parasite specific IgG1 ( D ) and IgG2c ( E ) production on day 21, n = 12, data shown is dilution 1:120. Values shown are means, unpaired T test. See also Figs S2 and S3 .

    Article Snippet: Parasite specific antibody was measured using biotinylated IgG1 or IgG2c (BD Biosciences).

    Techniques: Infection, Expressing, Real-time Polymerase Chain Reaction, MANN-WHITNEY

    Conditional deletion of bmal1 in dendritic cells abolishes diurnal variation in immune response to infection. Mice were generated which lacked bmal1 in CD11c positive cells (CD11c-bmal −/− ). ( A ) Bone marrow derived dendritic cells cultured from CD11c-bmal −/− mice on a PER2::luc background and their wildtype counterparts (bmal fl/fl ), were placed under photonmultiplier tubes (PMT) to monitor real-time luciferase activity as a readout of PER2 expression. ( B ) CD11c-bmal −/− and bmal fl/fl littermates were infected with T . muris at either ZT0 or ZT12. Worm burden (median presented) was assessed 21 days post infection, n = 15–16/group, combined data from two independent experiments. One way ANOVA, post hoc Tukey. ( C , D ) Parasite specific IgG2c and IgG1 production was measured on day 21. Serum was serially diluted and screened against parasite ES antigen (0.5 μg/ml); the data shown is dilution 1:160 only, as it falls within the linear range of the titration curve, n = 5–8, One Way ANOVA and post-hoc Tukey.

    Journal: Scientific Reports

    Article Title: The circadian regulator BMAL1 programmes responses to parasitic worm infection via a dendritic cell clock

    doi: 10.1038/s41598-018-22021-5

    Figure Lengend Snippet: Conditional deletion of bmal1 in dendritic cells abolishes diurnal variation in immune response to infection. Mice were generated which lacked bmal1 in CD11c positive cells (CD11c-bmal −/− ). ( A ) Bone marrow derived dendritic cells cultured from CD11c-bmal −/− mice on a PER2::luc background and their wildtype counterparts (bmal fl/fl ), were placed under photonmultiplier tubes (PMT) to monitor real-time luciferase activity as a readout of PER2 expression. ( B ) CD11c-bmal −/− and bmal fl/fl littermates were infected with T . muris at either ZT0 or ZT12. Worm burden (median presented) was assessed 21 days post infection, n = 15–16/group, combined data from two independent experiments. One way ANOVA, post hoc Tukey. ( C , D ) Parasite specific IgG2c and IgG1 production was measured on day 21. Serum was serially diluted and screened against parasite ES antigen (0.5 μg/ml); the data shown is dilution 1:160 only, as it falls within the linear range of the titration curve, n = 5–8, One Way ANOVA and post-hoc Tukey.

    Article Snippet: Parasite specific antibody was measured using biotinylated IgG1 or IgG2c (BD Biosciences).

    Techniques: Infection, Mouse Assay, Generated, Derivative Assay, Cell Culture, Luciferase, Activity Assay, Expressing, Titration

    Time of infection with Trichuris muris affects kinetics of expulsion. ( A ) The kinetics of expulsion of T . muris from the C57BL/6 mouse host. Worms establish in all mouse strains prior to expulsion (Establishment). The normal time course of expulsion, mediated by a Th2 response, is shown in blue and delayed expulsion by the red line (Expulsion). In C57 strains, expulsion is complete by approximately Day 30 (Resolution). ( B ) C57BL/6 mice were infected with 200 T . muris eggs at ZT0 or ZT12, and sacrificed at day 21 or 28, and worm burden accessed in the colon and caecum, combined data from 2 independent experiments, Mann Whitney tests. ( C , D ) Parasite-specific IgG1 (n = 6) and IgG2c (n = 10) production on days 21 and 28 respectively. Serum was serially diluted and screened against parasite ES antigen (0.5 μg/ml); data shown is 1:320 dilution, Mann Whitney tests. ( E ) Total IgE (day 21; n = 8 per group). ( F ) Mucosal mast cell protease-1 (MCPT-1; day 21; n = 8 per group) Mann Whitney tests. Mesenteric lymph node cell (MLN) IL-13 ( G ) and IFNγ ( H ) profiles at day 21post infection either at ZT0 or ZT12 cultured with T . muris E/S (50 µg/ml) for 48 h. Supernatants analysed using cytometric bead array (CBA), n = 8. Maximum level of detection for IFNγ 3000 pg/ml. Mann Whitney tests; data is representative of 2 repeats. All bars represent median. ( I ) Quantification of macrophages and dendritic cells in the large intestinal lamina propria by flow cytometry 0–3 days post infection, n = 3; Kruskal-Wallis test. Immunohistochemical staining for macrophages ( J ) (F4/80) and dendritic cells ( K ) (CD11c) in gut tissue on the day of or 1 day post infection. Positive cells are stained with DAB (Brown), hameatoxylin (blue) counterstain, scale bar represents 100 μm. See also Fig. S1 .

    Journal: Scientific Reports

    Article Title: The circadian regulator BMAL1 programmes responses to parasitic worm infection via a dendritic cell clock

    doi: 10.1038/s41598-018-22021-5

    Figure Lengend Snippet: Time of infection with Trichuris muris affects kinetics of expulsion. ( A ) The kinetics of expulsion of T . muris from the C57BL/6 mouse host. Worms establish in all mouse strains prior to expulsion (Establishment). The normal time course of expulsion, mediated by a Th2 response, is shown in blue and delayed expulsion by the red line (Expulsion). In C57 strains, expulsion is complete by approximately Day 30 (Resolution). ( B ) C57BL/6 mice were infected with 200 T . muris eggs at ZT0 or ZT12, and sacrificed at day 21 or 28, and worm burden accessed in the colon and caecum, combined data from 2 independent experiments, Mann Whitney tests. ( C , D ) Parasite-specific IgG1 (n = 6) and IgG2c (n = 10) production on days 21 and 28 respectively. Serum was serially diluted and screened against parasite ES antigen (0.5 μg/ml); data shown is 1:320 dilution, Mann Whitney tests. ( E ) Total IgE (day 21; n = 8 per group). ( F ) Mucosal mast cell protease-1 (MCPT-1; day 21; n = 8 per group) Mann Whitney tests. Mesenteric lymph node cell (MLN) IL-13 ( G ) and IFNγ ( H ) profiles at day 21post infection either at ZT0 or ZT12 cultured with T . muris E/S (50 µg/ml) for 48 h. Supernatants analysed using cytometric bead array (CBA), n = 8. Maximum level of detection for IFNγ 3000 pg/ml. Mann Whitney tests; data is representative of 2 repeats. All bars represent median. ( I ) Quantification of macrophages and dendritic cells in the large intestinal lamina propria by flow cytometry 0–3 days post infection, n = 3; Kruskal-Wallis test. Immunohistochemical staining for macrophages ( J ) (F4/80) and dendritic cells ( K ) (CD11c) in gut tissue on the day of or 1 day post infection. Positive cells are stained with DAB (Brown), hameatoxylin (blue) counterstain, scale bar represents 100 μm. See also Fig. S1 .

    Article Snippet: Parasite specific antibody was measured using biotinylated IgG1 or IgG2c (BD Biosciences).

    Techniques: Infection, Mouse Assay, MANN-WHITNEY, Cell Culture, Crocin Bleaching Assay, Flow Cytometry, Cytometry, Immunohistochemistry, Staining

    Immunization with TLR ligand–containing adjuvants bypasses the requirement for ZBTB20. (A) ELISA measurements of serum NP-specific IgG1, IgG2c, and IgG2b titers from Zbtb20 +/+ or Zbtb20 trap/trap fetal liver chimeras at 2, 9, and 19 wk after immunization with TLR ligand–adjuvanted NP-CGG (error bars depict geometric means ± 95% confidence interval). Data are cumulative from two experiments with 9–14 Zbtb20 +/+ and 10–15 Zbtb20 trap/trap chimeric mice per time point. Statistical significance was determined by the Mann–Whitney test. ns, not significant (P > 0.05). (B) ELISPOT analysis of NP-specific IgG1 BM ASCs from Zbtb20 +/+ ( n = 7) or Zbtb20 trap/trap ( n = 9) fetal liver chimeras at 20 wk after immunization (mean values ± SEM). Statistical significance was determined with an unpaired Student’s two-tailed t test. ns, not significant (P > 0.05). Data are cumulative from two independent experiments. (C) ELISA measurements of serum WNV-specific IgG titers from Zbtb20 +/+ ( n = 3) or Zbtb20 trap/trap ( n = 5) fetal liver chimeras at 21 wk after vaccination (error bars depict geometric means ± 95% confidence interval). Statistical significance was determined by the Mann–Whitney test. ns, not significant (P > 0.05). (D) ELISPOT assays of WNV-specific ASCs. WNV E protein–specific IgG-secreting cells in the BM from Zbtb20 +/+ ( n = 3) or Zbtb20 trap/trap ( n = 6) chimeric mice were quantified at 26 wk after vaccination. Mean values ± SEM are shown. Statistical significance was determined with an unpaired Student’s two-tailed t test. (C and D) Data are representative of two experiments, each with three to six mice per genotype.

    Journal: The Journal of Experimental Medicine

    Article Title: Adjuvant-specific regulation of long-term antibody responses by ZBTB20

    doi: 10.1084/jem.20131821

    Figure Lengend Snippet: Immunization with TLR ligand–containing adjuvants bypasses the requirement for ZBTB20. (A) ELISA measurements of serum NP-specific IgG1, IgG2c, and IgG2b titers from Zbtb20 +/+ or Zbtb20 trap/trap fetal liver chimeras at 2, 9, and 19 wk after immunization with TLR ligand–adjuvanted NP-CGG (error bars depict geometric means ± 95% confidence interval). Data are cumulative from two experiments with 9–14 Zbtb20 +/+ and 10–15 Zbtb20 trap/trap chimeric mice per time point. Statistical significance was determined by the Mann–Whitney test. ns, not significant (P > 0.05). (B) ELISPOT analysis of NP-specific IgG1 BM ASCs from Zbtb20 +/+ ( n = 7) or Zbtb20 trap/trap ( n = 9) fetal liver chimeras at 20 wk after immunization (mean values ± SEM). Statistical significance was determined with an unpaired Student’s two-tailed t test. ns, not significant (P > 0.05). Data are cumulative from two independent experiments. (C) ELISA measurements of serum WNV-specific IgG titers from Zbtb20 +/+ ( n = 3) or Zbtb20 trap/trap ( n = 5) fetal liver chimeras at 21 wk after vaccination (error bars depict geometric means ± 95% confidence interval). Statistical significance was determined by the Mann–Whitney test. ns, not significant (P > 0.05). (D) ELISPOT assays of WNV-specific ASCs. WNV E protein–specific IgG-secreting cells in the BM from Zbtb20 +/+ ( n = 3) or Zbtb20 trap/trap ( n = 6) chimeric mice were quantified at 26 wk after vaccination. Mean values ± SEM are shown. Statistical significance was determined with an unpaired Student’s two-tailed t test. (C and D) Data are representative of two experiments, each with three to six mice per genotype.

    Article Snippet: Wells were stained with biotinylated anti-IgG1A (10.9), anti-IgG1B (B68-2), anti-IgG1 (nonallotype specific; A85-1), anti-IgG2c (5.7), or anti-IgG2b (R12-3), purchased from BD.

    Techniques: Enzyme-linked Immunosorbent Assay, Mouse Assay, MANN-WHITNEY, Enzyme-linked Immunospot, Two Tailed Test

    IFN-γ–mediated expression of T-bet promotes IgG2c CSR but is not required for spontaneous GC formation. (A) Representative histograms of intracellular T-bet staining of WT B cells stimulated with R848, anti-IgM, anti-CD40 with or without exogenous IFN-β or IFN-γ. The gray histogram indicates unstimulated WT B cells. (B) T-bet mean fluorescence intensity (MFI) in stimulated B cells. (A and B) Data are representative of two or more independent experiments. (C, left) Representative histogram of Was −/− chimera intranuclear T-bet expression in GC (PNA + GL7 + ) versus naive (non-GC; PNA − GL7 − ) WT B cells. The gray histogram indicates Tbx21 −/− B cells. (Right) T-bet mean fluorescence intensity in total B cells from WT ( n = 2) and Was −/− ( n = 7) chimeras, as well as naive and GC B cells from Was −/− chimeras. (D) Anti-dsDNA (left) and Sm/RNP (right) isotype and subclass-specific auto-Abs (normalized to mean Was −/− chimera titer). (E–H) Spleen weight (E) and number of splenic CD4 + T cells (F), PNA + FAS + GC B cells (G), and CXCR5 + PD1 + Tfh cells (H). (I) Representative splenic sections stained with B220, PNA, and CD3 demonstrating PNA + GCs in both Was −/− and B cell–intrinsic Tbx21 −/− .Was −/− chimeras. Bars, 100 µm. (D–H) Data are representative of nine WT ( n = 16), nine Was −/− ( n = 34), and three B cell–intrinsic Was −/− .Tbx21 −/− ( n = 11) independent chimeras sacrificed at 24 wk. Error bars indicate SEM. *, P

    Journal: The Journal of Experimental Medicine

    Article Title: B cell IFN-γ receptor signaling promotes autoimmune germinal centers via cell-intrinsic induction of BCL-6

    doi: 10.1084/jem.20151724

    Figure Lengend Snippet: IFN-γ–mediated expression of T-bet promotes IgG2c CSR but is not required for spontaneous GC formation. (A) Representative histograms of intracellular T-bet staining of WT B cells stimulated with R848, anti-IgM, anti-CD40 with or without exogenous IFN-β or IFN-γ. The gray histogram indicates unstimulated WT B cells. (B) T-bet mean fluorescence intensity (MFI) in stimulated B cells. (A and B) Data are representative of two or more independent experiments. (C, left) Representative histogram of Was −/− chimera intranuclear T-bet expression in GC (PNA + GL7 + ) versus naive (non-GC; PNA − GL7 − ) WT B cells. The gray histogram indicates Tbx21 −/− B cells. (Right) T-bet mean fluorescence intensity in total B cells from WT ( n = 2) and Was −/− ( n = 7) chimeras, as well as naive and GC B cells from Was −/− chimeras. (D) Anti-dsDNA (left) and Sm/RNP (right) isotype and subclass-specific auto-Abs (normalized to mean Was −/− chimera titer). (E–H) Spleen weight (E) and number of splenic CD4 + T cells (F), PNA + FAS + GC B cells (G), and CXCR5 + PD1 + Tfh cells (H). (I) Representative splenic sections stained with B220, PNA, and CD3 demonstrating PNA + GCs in both Was −/− and B cell–intrinsic Tbx21 −/− .Was −/− chimeras. Bars, 100 µm. (D–H) Data are representative of nine WT ( n = 16), nine Was −/− ( n = 34), and three B cell–intrinsic Was −/− .Tbx21 −/− ( n = 11) independent chimeras sacrificed at 24 wk. Error bars indicate SEM. *, P

    Article Snippet: Plates were blocked for 1 h with 1% BSA in PBS before the addition of diluted serum for 2 h. Specific Abs were detected using goat anti–mouse IgM-, IgG-, or IgG2c-HRP (1:2,000 dilution; SouthernBiotech), and peroxidase reactions were developed using a OptEIA TMB substrate (BD).

    Techniques: Expressing, Staining, Fluorescence

    B cell antigen presentation initiates systemic autoimmunity. (A and B) Anti-dsDNA (A) and anti-Sm/RNP (B) IgM, IgG, and IgG2c auto-Abs at 12 (A) and 24 (B) wk after transplantation. (C and D) Spleen weight (C) and the number of splenic CD4 + T cells, CD11b + GR1 lo monocytes/macrophages, and CD11b + GR1 + neutrophils (D). (E) Representative FACS plots (gated on CD4 + T cells) showing naive (CD44 LO CD62L HI ) and EM (CD44 HI CD62L LO/HI ) CD4 + T cells. Numbers indicate the percentages within the CD44 LO CD62L HI and CD44 HI CD62L LO/HI gates. (F and G) Number of naive and EM CD4 + T cells (F) and PD1 + CXCR5 + Tfh cells (G). (H) Representative FACS plots (gated on B220 + ) and percentage of splenic PNA + FAS + GC B cells. (I) Proportion of PNA + FAS + GC B cells within the DZ (CXCR4 hi CD86 lo ) and light zone (LZ; CXCR4 lo CD86 hi ). Data are representative of two independent WT ( n = 6), Was −/− ( n = 8), and B cell MHCII −/− .Was −/− ( n = 8) chimeras. Error bars indicate SEM. *, P

    Journal: The Journal of Experimental Medicine

    Article Title: B cell IFN-γ receptor signaling promotes autoimmune germinal centers via cell-intrinsic induction of BCL-6

    doi: 10.1084/jem.20151724

    Figure Lengend Snippet: B cell antigen presentation initiates systemic autoimmunity. (A and B) Anti-dsDNA (A) and anti-Sm/RNP (B) IgM, IgG, and IgG2c auto-Abs at 12 (A) and 24 (B) wk after transplantation. (C and D) Spleen weight (C) and the number of splenic CD4 + T cells, CD11b + GR1 lo monocytes/macrophages, and CD11b + GR1 + neutrophils (D). (E) Representative FACS plots (gated on CD4 + T cells) showing naive (CD44 LO CD62L HI ) and EM (CD44 HI CD62L LO/HI ) CD4 + T cells. Numbers indicate the percentages within the CD44 LO CD62L HI and CD44 HI CD62L LO/HI gates. (F and G) Number of naive and EM CD4 + T cells (F) and PD1 + CXCR5 + Tfh cells (G). (H) Representative FACS plots (gated on B220 + ) and percentage of splenic PNA + FAS + GC B cells. (I) Proportion of PNA + FAS + GC B cells within the DZ (CXCR4 hi CD86 lo ) and light zone (LZ; CXCR4 lo CD86 hi ). Data are representative of two independent WT ( n = 6), Was −/− ( n = 8), and B cell MHCII −/− .Was −/− ( n = 8) chimeras. Error bars indicate SEM. *, P

    Article Snippet: Plates were blocked for 1 h with 1% BSA in PBS before the addition of diluted serum for 2 h. Specific Abs were detected using goat anti–mouse IgM-, IgG-, or IgG2c-HRP (1:2,000 dilution; SouthernBiotech), and peroxidase reactions were developed using a OptEIA TMB substrate (BD).

    Techniques: Transplantation Assay, FACS

    B cell–intrinsic IFN-γR signals promote spontaneous autoimmune GCs. (A) Number of IFN-γ + CD4 + T cells in WT, Was −/− , and B cell–intrinsic Tlr7 −/− , Tlr9 −/− , and MhcII −/− chimeras. (B) Anti-dsDNA and anti-Sm/RNP IgM, IgG, and IgG2c auto-Abs (12 wk after transplantation) in WT and Was −/− chimeras as well as Was −/− chimeras with global or B cell–intrinsic deletion of IFN-γR. (C) Spleen weight. (D) Number of splenic CD4 + T cells, CD11b + GR1 lo monocyte/macrophages, and CD11b + GR1 + neutrophils. (E and F) Representative FACS plots (E; gated on CD19 + ) and percentage (F) of splenic PNA + FAS + GC B cells. (G) Representative splenic sections stained with B220, PNA, and CD3. Bars, 100 µm. (H and I) Representative FACS plots (H; gated on CD4 + ) and number (I) of splenic PD1 + CXCR5 + Tfh cells. (J) Number of naive (CD44 LO CD62L HI ) and EM (CD44 HI CD62L LO/HI ) CD4 + T cells. (K) Total serum IgM, IgG, IgG2b, and IgG2c titers in the indicated chimeras. Data are representative of five WT ( n = 10), five Was −/− ( n = 18), two global Ifngr −/− Was −/− ( n = 10), and four B cell–intrinsic Ifngr −/− Was −/− ( n = 13) chimeras, as well as three Was −/− .Tlr7 −/− ( n = 14), three Was −/− .Tlr9 −/− ( n = 15), and two Was −/− .MhcII −/− ( n = 8) chimeras sacrificed at 24 wk after transplantation. Error bars indicate SEM. *, P

    Journal: The Journal of Experimental Medicine

    Article Title: B cell IFN-γ receptor signaling promotes autoimmune germinal centers via cell-intrinsic induction of BCL-6

    doi: 10.1084/jem.20151724

    Figure Lengend Snippet: B cell–intrinsic IFN-γR signals promote spontaneous autoimmune GCs. (A) Number of IFN-γ + CD4 + T cells in WT, Was −/− , and B cell–intrinsic Tlr7 −/− , Tlr9 −/− , and MhcII −/− chimeras. (B) Anti-dsDNA and anti-Sm/RNP IgM, IgG, and IgG2c auto-Abs (12 wk after transplantation) in WT and Was −/− chimeras as well as Was −/− chimeras with global or B cell–intrinsic deletion of IFN-γR. (C) Spleen weight. (D) Number of splenic CD4 + T cells, CD11b + GR1 lo monocyte/macrophages, and CD11b + GR1 + neutrophils. (E and F) Representative FACS plots (E; gated on CD19 + ) and percentage (F) of splenic PNA + FAS + GC B cells. (G) Representative splenic sections stained with B220, PNA, and CD3. Bars, 100 µm. (H and I) Representative FACS plots (H; gated on CD4 + ) and number (I) of splenic PD1 + CXCR5 + Tfh cells. (J) Number of naive (CD44 LO CD62L HI ) and EM (CD44 HI CD62L LO/HI ) CD4 + T cells. (K) Total serum IgM, IgG, IgG2b, and IgG2c titers in the indicated chimeras. Data are representative of five WT ( n = 10), five Was −/− ( n = 18), two global Ifngr −/− Was −/− ( n = 10), and four B cell–intrinsic Ifngr −/− Was −/− ( n = 13) chimeras, as well as three Was −/− .Tlr7 −/− ( n = 14), three Was −/− .Tlr9 −/− ( n = 15), and two Was −/− .MhcII −/− ( n = 8) chimeras sacrificed at 24 wk after transplantation. Error bars indicate SEM. *, P

    Article Snippet: Plates were blocked for 1 h with 1% BSA in PBS before the addition of diluted serum for 2 h. Specific Abs were detected using goat anti–mouse IgM-, IgG-, or IgG2c-HRP (1:2,000 dilution; SouthernBiotech), and peroxidase reactions were developed using a OptEIA TMB substrate (BD).

    Techniques: Transplantation Assay, FACS, Staining

    B cell–intrinsic IFNAR signals are not required for the development of humoral autoimmunity. (A) IFN-stimulated gene mRNA transcripts (fold change vs. WT chimera) in splenocytes from two or more independent chimeras. (B) Total splenic plasmacytoid DCs (pDCs) in WT ( n = 5) and Was −/− ( n = 9) mice from three independent chimeras. (C) Representative histograms. (Left) Gating of proliferated versus unproliferated subsets (assessed by Cell Trace dilution) in WT B cells stimulated with anti-IgM, R848, and IFN-β. (Middle) Relative surface PNA binding. (Right) AID expression in proliferated (red) versus unproliferated (blue) B cells. The gray histogram indicates unstimulated B cells. (D and E) Proliferation (D) and PNA binding (E) in splenic WT and Ifnar −/− B cells stimulated with the indicated combinations of anti-IgM, R848, anti-CD40, and IFN-β. (F) Tlr7 mRNA transcripts in sorted WT versus Ifnar −/− follicular mature (FM) and marginal zone (MZ) B cells (fold change vs. WT FM; Tlr7 −/− is shown as a negative control). (G and H) Anti-dsDNA (G) and anti-Sm/RNP (H) IgG and IgG2c auto-Ab in WT (white), Was −/− (black), and B cell Ifnar −/− Was −/− (gray) chimeras at 12 (G) and 24 (H) wk after transplantation. (I) Spleen weights. (J) Splenic CD4 + T cells and CD11b + GR1 lo monocytes/macrophages. (K) Representative FACS plots (left) and total number (right) of splenic PNA + FAS + GC B cells. (L) Representative splenic sections stained with B220, PNA, and CD3. (M) Number of splenic naive (CD44 LO CD62L HI ) and EM (CD44 HI CD62L LO/HI ) CD4 + T cells and PD1 + CXCR5 + Tfh cells. (N, left) Representative glomerular IF staining. (Right) Intensity of glomerular IgG, IgG2c, and C3 staining scored from 0 to 3 by two independent blinded observers. (A, B, and G–N) Data are representative of four independent WT ( n = 7), Was −/− ( n = 18), and B cell Ifnar −/− Was −/− ( n = 19) chimeras sacrificed at 24 wk after transplantation. (C–F) Data are representative of two or more independent experiments. Error bars indicate SEM. *, P

    Journal: The Journal of Experimental Medicine

    Article Title: B cell IFN-γ receptor signaling promotes autoimmune germinal centers via cell-intrinsic induction of BCL-6

    doi: 10.1084/jem.20151724

    Figure Lengend Snippet: B cell–intrinsic IFNAR signals are not required for the development of humoral autoimmunity. (A) IFN-stimulated gene mRNA transcripts (fold change vs. WT chimera) in splenocytes from two or more independent chimeras. (B) Total splenic plasmacytoid DCs (pDCs) in WT ( n = 5) and Was −/− ( n = 9) mice from three independent chimeras. (C) Representative histograms. (Left) Gating of proliferated versus unproliferated subsets (assessed by Cell Trace dilution) in WT B cells stimulated with anti-IgM, R848, and IFN-β. (Middle) Relative surface PNA binding. (Right) AID expression in proliferated (red) versus unproliferated (blue) B cells. The gray histogram indicates unstimulated B cells. (D and E) Proliferation (D) and PNA binding (E) in splenic WT and Ifnar −/− B cells stimulated with the indicated combinations of anti-IgM, R848, anti-CD40, and IFN-β. (F) Tlr7 mRNA transcripts in sorted WT versus Ifnar −/− follicular mature (FM) and marginal zone (MZ) B cells (fold change vs. WT FM; Tlr7 −/− is shown as a negative control). (G and H) Anti-dsDNA (G) and anti-Sm/RNP (H) IgG and IgG2c auto-Ab in WT (white), Was −/− (black), and B cell Ifnar −/− Was −/− (gray) chimeras at 12 (G) and 24 (H) wk after transplantation. (I) Spleen weights. (J) Splenic CD4 + T cells and CD11b + GR1 lo monocytes/macrophages. (K) Representative FACS plots (left) and total number (right) of splenic PNA + FAS + GC B cells. (L) Representative splenic sections stained with B220, PNA, and CD3. (M) Number of splenic naive (CD44 LO CD62L HI ) and EM (CD44 HI CD62L LO/HI ) CD4 + T cells and PD1 + CXCR5 + Tfh cells. (N, left) Representative glomerular IF staining. (Right) Intensity of glomerular IgG, IgG2c, and C3 staining scored from 0 to 3 by two independent blinded observers. (A, B, and G–N) Data are representative of four independent WT ( n = 7), Was −/− ( n = 18), and B cell Ifnar −/− Was −/− ( n = 19) chimeras sacrificed at 24 wk after transplantation. (C–F) Data are representative of two or more independent experiments. Error bars indicate SEM. *, P

    Article Snippet: Plates were blocked for 1 h with 1% BSA in PBS before the addition of diluted serum for 2 h. Specific Abs were detected using goat anti–mouse IgM-, IgG-, or IgG2c-HRP (1:2,000 dilution; SouthernBiotech), and peroxidase reactions were developed using a OptEIA TMB substrate (BD).

    Techniques: Mouse Assay, Binding Assay, Expressing, Negative Control, Transplantation Assay, FACS, Staining

    Tfr cells regulate the production of anti-SRBC IgG1 and anti-SRBC IgG2b. Control and Stat3FC mice were immunized with SRBC via i.p. injection. Serum samples were collected at 25 dpi. Anti-SRBC IgM, IgG1, IgG2b, IgG2c, IgG3 and IgA titers are shown. SRBC-specific IgD and IgE were not detectable in the serum. The X-axis shows the dilution factors. Graphs show mean +/- SEM, n = 4. ** p

    Journal: PLoS ONE

    Article Title: Stat3 Is Important for Follicular Regulatory T Cell Differentiation

    doi: 10.1371/journal.pone.0155040

    Figure Lengend Snippet: Tfr cells regulate the production of anti-SRBC IgG1 and anti-SRBC IgG2b. Control and Stat3FC mice were immunized with SRBC via i.p. injection. Serum samples were collected at 25 dpi. Anti-SRBC IgM, IgG1, IgG2b, IgG2c, IgG3 and IgA titers are shown. SRBC-specific IgD and IgE were not detectable in the serum. The X-axis shows the dilution factors. Graphs show mean +/- SEM, n = 4. ** p

    Article Snippet: For analysis of IgG1 (A85-1, BD Biosciences), IgG2b (R12-3, BD Biosciences), IgG2c (Abcam) and IgG3 (R40-82, BD Biosciences), biotin-labeled secondary Abs were used along with Avidin-HRP (BD Biosciences).

    Techniques: Mouse Assay, Injection

    Basal titers of serum IgM, IgG1, IgG2b, IgG2c and IgG3 are not altered in Stat3FC mice. Serum samples from unimmunized control and Stat3FC mice were collected. IgM, IgG1, IgG2b, IgG2c and IgG3 titers are shown. The X-axis shows the dilution factors. Graphs show mean +/- SEM, n = 3. No significant differences in basal Ig isotype levels were observed by two-way ANOVA. Data are representative of two independent experiments with similar results.

    Journal: PLoS ONE

    Article Title: Stat3 Is Important for Follicular Regulatory T Cell Differentiation

    doi: 10.1371/journal.pone.0155040

    Figure Lengend Snippet: Basal titers of serum IgM, IgG1, IgG2b, IgG2c and IgG3 are not altered in Stat3FC mice. Serum samples from unimmunized control and Stat3FC mice were collected. IgM, IgG1, IgG2b, IgG2c and IgG3 titers are shown. The X-axis shows the dilution factors. Graphs show mean +/- SEM, n = 3. No significant differences in basal Ig isotype levels were observed by two-way ANOVA. Data are representative of two independent experiments with similar results.

    Article Snippet: For analysis of IgG1 (A85-1, BD Biosciences), IgG2b (R12-3, BD Biosciences), IgG2c (Abcam) and IgG3 (R40-82, BD Biosciences), biotin-labeled secondary Abs were used along with Avidin-HRP (BD Biosciences).

    Techniques: Mouse Assay

    CTLA-4 blockade amplifies serum HIV Gag- and Env-specific antibody responses. Post-prime serum was collected 13 days after the first VLP immunization; Post-boost 1 serum was collected 11–13 days after the second VLP immunization. Post-prime and Post-boost 1 time points analyzed serum from 10 mice per immunization group from Cohorts 1 and cohorts 2. The Post-boost 2 time point analyzed serum from the 15 mice (5 per group) in Cohort 2 and was collected 7 days after the third VLP immunization. ( A ) Total Env-specific IgG and ( B ) total Gag-specific IgG were analyzed through ELISA. Quantitative ELISAs with standard curves for IgG1, IgG2b, IgG2c, and IgG3 were performed on all serum samples from Cohorts 1 and 2 to detect HIV Env- and Gag-specific IgG subtypes. Shown at each of the following time points: ( C ) post-prime, ( D ) Env post-prime Gag, ( E ) post-boost 1 Env, ( F ) post-boost 1 Gag, ( G ) post-boost 2 Env, and ( H ) post-boost 2 Gag IgG subtypes. The results above are the cumulative results of two independent experiments. P-values were determined by one-way ANOVA and Tukey post-hoc analysis for multiple comparisons. A line on top of two groups indicates statistical differences between these two groups. **** p

    Journal: Vaccines

    Article Title: CTLA-4 Blockade, during HIV Virus-Like Particles Immunization, Alters HIV-Specific B-Cell Responses

    doi: 10.3390/vaccines8020284

    Figure Lengend Snippet: CTLA-4 blockade amplifies serum HIV Gag- and Env-specific antibody responses. Post-prime serum was collected 13 days after the first VLP immunization; Post-boost 1 serum was collected 11–13 days after the second VLP immunization. Post-prime and Post-boost 1 time points analyzed serum from 10 mice per immunization group from Cohorts 1 and cohorts 2. The Post-boost 2 time point analyzed serum from the 15 mice (5 per group) in Cohort 2 and was collected 7 days after the third VLP immunization. ( A ) Total Env-specific IgG and ( B ) total Gag-specific IgG were analyzed through ELISA. Quantitative ELISAs with standard curves for IgG1, IgG2b, IgG2c, and IgG3 were performed on all serum samples from Cohorts 1 and 2 to detect HIV Env- and Gag-specific IgG subtypes. Shown at each of the following time points: ( C ) post-prime, ( D ) Env post-prime Gag, ( E ) post-boost 1 Env, ( F ) post-boost 1 Gag, ( G ) post-boost 2 Env, and ( H ) post-boost 2 Gag IgG subtypes. The results above are the cumulative results of two independent experiments. P-values were determined by one-way ANOVA and Tukey post-hoc analysis for multiple comparisons. A line on top of two groups indicates statistical differences between these two groups. **** p

    Article Snippet: Quantitative IgG Subtype ELISAMicro-vinyl plates (96-well, Corning Costar, Corning, NY, USA) were coated with a standard curve of purified IgG1 (BD Pharminogen #554121, San Diego, CA, USA), IgG2b (eBioscience eBMG2b #14732-82, San Diego, CA, UAS), IgG2c (GeneTex #GTX35043, Irvine, CA, USA), or IgG3 (eBioscience #14-4742-82, San Diego, CA, USA) protein while remaining wells were coated with 0.5 μg/mL BaL gp120 recombinant protein (NIH AIDS reagent #4961, Germantown, MD, USA) in a sodium bicarbonate buffer and incubated overnight.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    c1(70–100) dKI mice show a specific breach of B cell anergy to ssDNA. Serum from 8M B6 (circles), c1(96–100) (squares) and c1(70–100) (triangles) WT (filled) and dKI (open) mice was diluted 1:100 and assessed by ELISA for production of (A) anti-ssDNA and (B) anti-dsDNA IgM, IgM a/b , IgG, IgG2a/IgG2a a and IgG2c/IgG2a b autoAbs. Scatterplots show data from multiple independent experiments with n = 3–16 mice each. Symbols represent individual mice and lines show the median. Kruskal-Wallis non-parametric tests with Dunn’s post-test were used to compare B6 with c1 animals within each genotype. *p

    Journal: PLoS ONE

    Article Title: Impaired B cell anergy is not sufficient to breach tolerance to nuclear antigen in Vκ8/3H9 lupus-prone mice

    doi: 10.1371/journal.pone.0236664

    Figure Lengend Snippet: c1(70–100) dKI mice show a specific breach of B cell anergy to ssDNA. Serum from 8M B6 (circles), c1(96–100) (squares) and c1(70–100) (triangles) WT (filled) and dKI (open) mice was diluted 1:100 and assessed by ELISA for production of (A) anti-ssDNA and (B) anti-dsDNA IgM, IgM a/b , IgG, IgG2a/IgG2a a and IgG2c/IgG2a b autoAbs. Scatterplots show data from multiple independent experiments with n = 3–16 mice each. Symbols represent individual mice and lines show the median. Kruskal-Wallis non-parametric tests with Dunn’s post-test were used to compare B6 with c1 animals within each genotype. *p

    Article Snippet: ELISAs Serum levels of anti-ssDNA or anti-dsDNA IgM, IgMa , IgMb , IgG, IgG2a, IgG2aa , IgG2ab , and IgG2c antibodies were measured by ELISA as previously described [ ].

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    Hemagglutinin (HA)- and neuraminidase (NA)-specific antibody responses in vivo . Levels of HA from Cal7-specific total IgG (A) , IgG1 (B) , and IgG2c (C) ; of NA from Cal7-specific total IgG (D) , IgG1 (E) , and IgG2c (F) ; and of HA from PR8-specific total IgG (G) , IgG1 (H) , and IgG2c (I) in plasma were evaluated by using ELISA 7 days after final immunization. The same plasma samples as used in Figure 4 were used here. We used 800- (•), 4000- (■), and 20,000- (▴) fold diluted plasma samples for (A–F) and 32- (•), 160- (■), and 800- (▴) fold diluted plasma samples for (G–I) . n = 5. Data are means ± SD. Significant differences were analyzed only in the 800-fold-diluted plasma samples (A–F) and the 32-fold-diluted plasma samples (G–I) . † P

    Journal: Frontiers in Immunology

    Article Title: Lipid Nanoparticles Potentiate CpG-Oligodeoxynucleotide-Based Vaccine for Influenza Virus

    doi: 10.3389/fimmu.2019.03018

    Figure Lengend Snippet: Hemagglutinin (HA)- and neuraminidase (NA)-specific antibody responses in vivo . Levels of HA from Cal7-specific total IgG (A) , IgG1 (B) , and IgG2c (C) ; of NA from Cal7-specific total IgG (D) , IgG1 (E) , and IgG2c (F) ; and of HA from PR8-specific total IgG (G) , IgG1 (H) , and IgG2c (I) in plasma were evaluated by using ELISA 7 days after final immunization. The same plasma samples as used in Figure 4 were used here. We used 800- (•), 4000- (■), and 20,000- (▴) fold diluted plasma samples for (A–F) and 32- (•), 160- (■), and 800- (▴) fold diluted plasma samples for (G–I) . n = 5. Data are means ± SD. Significant differences were analyzed only in the 800-fold-diluted plasma samples (A–F) and the 32-fold-diluted plasma samples (G–I) . † P

    Article Snippet: Both LNP-CpGs induced not only IgG2c specific to HA from Cal7 ( ) but also IgG2c specific to HA from PR8 , although the levels of IgG2c specific to HA from PR8 were lower than those of IgG2c specific to HA from Cal7.

    Techniques: In Vivo, Enzyme-linked Immunosorbent Assay

    Preventive effects against heterosubtypic influenza virus. Mice were immunized subcutaneously with SV from Tex50 alone, SV plus CpG ODN, or SV plus either LNP-CpG. (A–C) Antibody responses. Levels of total IgG (A) , IgG1 (B) , and IgG2c (C) specific to SV from Tex50 in plasma were evaluated by using ELISA 7 days after final immunization. We used 4000- (•), 20,000- (■), and 100,000- (▴) fold diluted plasma samples. (D,E) Preventive effects against heterosubtypic Cal7. Ten days after final immunization, mice were challenged with Cal7. Percentages of initial body weights (D) and survival rates (E) were monitored after challenge with Cal7. (A–E) n = 5. Data are means ± SD. (A–C) Significant differences were analyzed only in the 4,000-fold-diluted plasma samples. †† P

    Journal: Frontiers in Immunology

    Article Title: Lipid Nanoparticles Potentiate CpG-Oligodeoxynucleotide-Based Vaccine for Influenza Virus

    doi: 10.3389/fimmu.2019.03018

    Figure Lengend Snippet: Preventive effects against heterosubtypic influenza virus. Mice were immunized subcutaneously with SV from Tex50 alone, SV plus CpG ODN, or SV plus either LNP-CpG. (A–C) Antibody responses. Levels of total IgG (A) , IgG1 (B) , and IgG2c (C) specific to SV from Tex50 in plasma were evaluated by using ELISA 7 days after final immunization. We used 4000- (•), 20,000- (■), and 100,000- (▴) fold diluted plasma samples. (D,E) Preventive effects against heterosubtypic Cal7. Ten days after final immunization, mice were challenged with Cal7. Percentages of initial body weights (D) and survival rates (E) were monitored after challenge with Cal7. (A–E) n = 5. Data are means ± SD. (A–C) Significant differences were analyzed only in the 4,000-fold-diluted plasma samples. †† P

    Article Snippet: Both LNP-CpGs induced not only IgG2c specific to HA from Cal7 ( ) but also IgG2c specific to HA from PR8 , although the levels of IgG2c specific to HA from PR8 were lower than those of IgG2c specific to HA from Cal7.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    Influenza-virus-specific antibody responses in vivo . Mice were immunized subcutaneously with split vaccine (SV) alone, SV plus CpG ODN, SV plus either LNP-CpG, or SV plus alum. (A–F) Antibody responses. Levels of SV-specific total IgG (A) , IgG1 (B) , and IgG2c (C) , and of PR8-specific total IgG (D) , IgG1 (E) , and IgG2c (F) in plasma were evaluated by using ELISA 7 days after final immunization. We used 6000- (•), 30,000- (■), and 150,000- (▴) fold diluted plasma samples. (G,H) Neutralization titers against influenza virus. Neutralization titers in plasma samples against Cal7 (G) and PR8 (H) were evaluated. (A–F) n = 5 per group. Data are means ± SD. Significant differences were analyzed only in the 6,000-fold-diluted plasma samples. † P

    Journal: Frontiers in Immunology

    Article Title: Lipid Nanoparticles Potentiate CpG-Oligodeoxynucleotide-Based Vaccine for Influenza Virus

    doi: 10.3389/fimmu.2019.03018

    Figure Lengend Snippet: Influenza-virus-specific antibody responses in vivo . Mice were immunized subcutaneously with split vaccine (SV) alone, SV plus CpG ODN, SV plus either LNP-CpG, or SV plus alum. (A–F) Antibody responses. Levels of SV-specific total IgG (A) , IgG1 (B) , and IgG2c (C) , and of PR8-specific total IgG (D) , IgG1 (E) , and IgG2c (F) in plasma were evaluated by using ELISA 7 days after final immunization. We used 6000- (•), 30,000- (■), and 150,000- (▴) fold diluted plasma samples. (G,H) Neutralization titers against influenza virus. Neutralization titers in plasma samples against Cal7 (G) and PR8 (H) were evaluated. (A–F) n = 5 per group. Data are means ± SD. Significant differences were analyzed only in the 6,000-fold-diluted plasma samples. † P

    Article Snippet: Both LNP-CpGs induced not only IgG2c specific to HA from Cal7 ( ) but also IgG2c specific to HA from PR8 , although the levels of IgG2c specific to HA from PR8 were lower than those of IgG2c specific to HA from Cal7.

    Techniques: In Vivo, Mouse Assay, Enzyme-linked Immunosorbent Assay, Neutralization

    rAd-mIL-28B induced the reduction of anti-Ag85B antibodies in mice primed with BCG and boosted with subunit vaccine. Mice were immunized according to . At week 31, the mice were bled to confirm levels of the anti-Ag85B antibodies IgG2c and IgG1

    Journal: International Immunology

    Article Title: IL-28B down-regulates regulatory T cells but does not improve the protective immunity following tuberculosis subunit vaccine immunization

    doi: 10.1093/intimm/dxv061

    Figure Lengend Snippet: rAd-mIL-28B induced the reduction of anti-Ag85B antibodies in mice primed with BCG and boosted with subunit vaccine. Mice were immunized according to . At week 31, the mice were bled to confirm levels of the anti-Ag85B antibodies IgG2c and IgG1

    Article Snippet: Subsequently, the plates were incubated with the polyclonal HRP-conjugated second antibodies rabbit anti-mouse IgG1 (Rockland, PA, USA) or goat anti-mouse IgG2c (Bethyl, AL, USA) for 1h at 37°C.

    Techniques: Mouse Assay

    rAd-mIL-28B treatment increased Ag85B-specific IgG1 and decreased Ag85B-specific IgG2c following EAMMH vaccination. The procedure of immunization was performed according to . At week 10, mice were bled to detect serum antibodies with indirect ELISA.

    Journal: International Immunology

    Article Title: IL-28B down-regulates regulatory T cells but does not improve the protective immunity following tuberculosis subunit vaccine immunization

    doi: 10.1093/intimm/dxv061

    Figure Lengend Snippet: rAd-mIL-28B treatment increased Ag85B-specific IgG1 and decreased Ag85B-specific IgG2c following EAMMH vaccination. The procedure of immunization was performed according to . At week 10, mice were bled to detect serum antibodies with indirect ELISA.

    Article Snippet: Subsequently, the plates were incubated with the polyclonal HRP-conjugated second antibodies rabbit anti-mouse IgG1 (Rockland, PA, USA) or goat anti-mouse IgG2c (Bethyl, AL, USA) for 1h at 37°C.

    Techniques: Mouse Assay, Indirect ELISA

    Serum IgG1 and IgG2a (A) and IgG2b and IgG2c (B) responses to rHag B in rats immunized s.c. with rHag B (group A) and in rats immunized s.c. with rHag B and orally challenged with freshly harvested P. gingivalis 33277 (group B). Values are the geometric means ×/÷ SD of antibody activity in serum samples from six rats/group. The results are from one experiment but representative of two separate experiments.

    Journal: Infection and Immunity

    Article Title: Host Responses to Recombinant Hemagglutinin B of Porphyromonas gingivalis in an Experimental Rat Model

    doi:

    Figure Lengend Snippet: Serum IgG1 and IgG2a (A) and IgG2b and IgG2c (B) responses to rHag B in rats immunized s.c. with rHag B (group A) and in rats immunized s.c. with rHag B and orally challenged with freshly harvested P. gingivalis 33277 (group B). Values are the geometric means ×/÷ SD of antibody activity in serum samples from six rats/group. The results are from one experiment but representative of two separate experiments.

    Article Snippet: Briefly, individual wells of flat-bottom 96-well plates (Nalge Nunc International, Roshilde, Denmark) were coated with rHag B or with P. gingivalis 33277 WC prepared in bicarbonate-carbonate buffer (pH 9.6; 100 μl; 1 μg of rHag B per ml or 5 × 108 cells/ml) or with anti-rat α, μ, or γ heavy-chain antibody (UAB Immunochemical Core Facility) or affinity purified anti-rat IgG1, IgG2a, IgG2b, or IgG2c (The Binding Site Inc., San Diego, Calif.) in borate-buffered saline (pH 8.2).

    Techniques: Activity Assay

    Effect of nasal administration of pCol (24-38) in groups of WKY rats ( n = 5 to 6) after the onset of disease on levels of circulating IgG1 ( A ), IgG2a ( B ), IgG2b ( C ), and IgG2c ( D ) anti-α3(IV)NC1 antibody concentrations. Results shown represent the mean ± SD of each group at day 28 after immunization.

    Journal: The American Journal of Pathology

    Article Title: Mucosal Tolerance Induced by an Immunodominant Peptide from Rat ?3(IV)NC1 in Established Experimental Autoimmune Glomerulonephritis

    doi: 10.2353/ajpath.2009.081041

    Figure Lengend Snippet: Effect of nasal administration of pCol (24-38) in groups of WKY rats ( n = 5 to 6) after the onset of disease on levels of circulating IgG1 ( A ), IgG2a ( B ), IgG2b ( C ), and IgG2c ( D ) anti-α3(IV)NC1 antibody concentrations. Results shown represent the mean ± SD of each group at day 28 after immunization.

    Article Snippet: The isotypes of circulating anti-α3(IV)NC1 antibodies were detected by mouse monoclonal antibodies specific for rat IgG1 and IgG2a IgG2b and IgG2c (Serotec Ltd.), followed by goat anti-mouse, IgG (Serotec Ltd.).

    Techniques: