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  • 99
    Vector Laboratories biotinylated anti mouse
    Id1 is expressed in ECs and ST fibroblasts. a mRNA was isolated from HMVECs and fibroblasts were isolated from NL and RA ST. mRNA was reverse transcribed into cDNA and underwent PCR for 40 cycles. RA fibroblasts showed significantly elevated Id1 expression compared to NL ST fibroblasts and HMVECs. b Id1 is expressed in RA STs. IHC was performed on RA, OA, and NL ST cryosections. Tissues were blocked and then incubated using a mouse anti-human Id1 (Abcam) primary antibody. After washing, tissues were incubated with a <t>biotinylated</t> anti-mouse secondary antibody (Vector Laboratories). Tissues were washed and subsequently developed with the Vectastain ABC kit (Vector Laboratories). Id1 is found on synovial cells (SNC) in the RA ST. c For immunofluorescence staining, RA, osteoarthritis (OA), and normal (NL) ST sections were fixed in cold acetone for 30 min. The STs were blocked with 5 % donkey serum and 20 % fetal bovine serum (FBS) in PBS at 37 °C for 1 h, and then incubated with rabbit anti-human Id1 antibody (Abcam, 10 μg/ml) or purified nonspecific rabbit IgG for 1 h at 37 °C in blocking buffer. The synovial tissues samples were washed with PBS, and a 1:200 dilution in blocking buffer of fluorescent-conjugated donkey anti-rabbit antibody was added and incubated for an additional 1 h at 37 °C. As shown, we could validate Id1 staining in RA ST similar to what was found using IHC (×400). FLS fibroblast-like synoviocytes, HMVEC human dermal microvascular endothelial cell, Id1 inhibitor of DNA binding 1, IgG immunoglobulin, NL normal, RA rheumatoid arthritis, SNC synovial cell
    Biotinylated Anti Mouse, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 99/100, based on 2573 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher anti mouse secondary antibody
    Id1 is expressed in ECs and ST fibroblasts. a mRNA was isolated from HMVECs and fibroblasts were isolated from NL and RA ST. mRNA was reverse transcribed into cDNA and underwent PCR for 40 cycles. RA fibroblasts showed significantly elevated Id1 expression compared to NL ST fibroblasts and HMVECs. b Id1 is expressed in RA STs. IHC was performed on RA, OA, and NL ST cryosections. Tissues were blocked and then incubated using a mouse anti-human Id1 (Abcam) primary antibody. After washing, tissues were incubated with a <t>biotinylated</t> anti-mouse secondary antibody (Vector Laboratories). Tissues were washed and subsequently developed with the Vectastain ABC kit (Vector Laboratories). Id1 is found on synovial cells (SNC) in the RA ST. c For immunofluorescence staining, RA, osteoarthritis (OA), and normal (NL) ST sections were fixed in cold acetone for 30 min. The STs were blocked with 5 % donkey serum and 20 % fetal bovine serum (FBS) in PBS at 37 °C for 1 h, and then incubated with rabbit anti-human Id1 antibody (Abcam, 10 μg/ml) or purified nonspecific rabbit IgG for 1 h at 37 °C in blocking buffer. The synovial tissues samples were washed with PBS, and a 1:200 dilution in blocking buffer of fluorescent-conjugated donkey anti-rabbit antibody was added and incubated for an additional 1 h at 37 °C. As shown, we could validate Id1 staining in RA ST similar to what was found using IHC (×400). FLS fibroblast-like synoviocytes, HMVEC human dermal microvascular endothelial cell, Id1 inhibitor of DNA binding 1, IgG immunoglobulin, NL normal, RA rheumatoid arthritis, SNC synovial cell
    Anti Mouse Secondary Antibody, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 2084 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Abcam goat anti rabbit igg h l hrp
    F-ATPase/VGCC protein complex verification. a. 1D immunoblot analysis of the VGCC α2δ-1 subunit in neutrophils. The VGCC α2δ-1 subunit-containing protein complexes in a NativePAGE Novex 4–16% Bis-Tris gel were detected at approximately 750 kDa via immunoblotting, the same position as the F-ATPase F1 β-subunit. b. 2D immunoblot analysis of the F-ATPase and VGCC protein complex. Plasma membrane lysed by 2% digitonin was loaded and run on a BN-PAGE gel. One lane (Sample) was cut and equilibrated in SDS loading buffer, followed by fixation on a SDS-PAGE gel for 2D electrophoretic analysis. Two separate immunoblot analyses with a primary antibody against the VGCC α2δ-1 subunit or the F-ATPase F1 β-subunit were performed. The F-ATPase F1 β-subunit appears vertically below the VGCC α2δ-1 subunit in the same PVDF membrane. c. Immunofluorescence colocalization analysis of F-ATPase and VGCC. Representative images show that the VGCC α2δ-1 subunit and F-ATPase F1 β-subunit colocalized on the plasma membrane of neutrophils. d. Immunoprecipitation of VGCC α2δ-1 subunit and F-ATPase from plasma membrane protein in human neutrophils. Samples were immunoprecipitated with a mouse monoclonal antibody against α2 of the VGCC α2δ-1 subunit. F-ATPase was detected with a mouse monoclonal antibody against the F-ATPase F1 α-subunit or β-subunit and an <t>HRP-conjugated</t> goat anti-mouse <t>IgG</t> light chain specifically bound antibody. IgG control experiment did not detect F-ATPase
    Goat Anti Rabbit Igg H L Hrp, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 2991 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Proteintech mouse gapdh monoclonal
    miR-137 represses cholangiocarcinoma cell proliferation in vitro . (A) Fluorescence microscope examination (magnification, ×200) and reverse transcription-quantitative PCR were used to detect the infection efficiency of miR-137 overexpression (LV-miR-137) and NC lentiviruses. (B) The effect of miR-137 on cholangiocarcinoma cell proliferation was detected by the Cell Counting Kit-8 assay. (C) The effect of miR-137 on cholangiocarcinoma cell colony formation ability was detected using colony formation assays. (D) Cell cycle distribution was analyzed after miR-137 overexpression in TFK-1 and HuCCT1 cells. (E) Western blotting was used to detect the expression of <t>CDK2</t> and cyclin D1 in the miR-137 overexpression and normal control groups. <t>GAPDH</t> was used as the loading control. * P
    Mouse Gapdh Monoclonal, supplied by Proteintech, used in various techniques. Bioz Stars score: 99/100, based on 1626 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Proteintech mouse beta actin monoclonal
    RNP purification from animal tissue. Mouse brain tissue was cryo-grinded and UV-irradiated before (Hot)-PTex was performed. Western blot against HuR (ELAVL1) demonstrates recovery of cross-linked HuR from mouse tissue while <t>beta-actin</t> (ACTB) is efficiently depleted. For full blots see Supplementary Figure 15
    Mouse Beta Actin Monoclonal, supplied by Proteintech, used in various techniques. Bioz Stars score: 99/100, based on 1613 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Bristol Myers ctla4 ig
    Signature of therapeutic response in TN-09. ( a ) Analysis of the 1509 probe sets regulated between the <t>CTLA4-Ig</t> responders ( n = 8) and rapidly progressing placebo-treated individuals ( n = 7) matched for baseline I . I . 359 at the 3 month time point. The top three panels (left to right) illustrate two-way clustering of the selected and excluded participants in each arm. Left panel, mean expression of the four groups. Middle panel, participants selected for the analysis. Right panel, remaining CTLA4-Ig- and placebo-treated participants. Lower three panels, expression levels of a selected set of well-annotated transcripts. The colour bar indicates assigned treatment arm: red, placebo; blue, CTLA4-Ig. Note: samples were not available for two placebo-treated participants at the 3 month time point. The annotated dataset is available from the corresponding author on request. ( b ) Mean expression levels of the 1509 differentially induced probe sets at 12 and 24 months. ( c ) Upstream regulator analysis using IPA. A z score > 2.0 is significantly activated; a z score > −2.0 is significantly inhibited. ( d ) CTLA4-Ig add-back experiment. The mean induced expression levels of the 1509 probe sets after plasma of five local untreated individuals with diabetes were supplemented with 0 μg/ml, 25 μg/ml or 82 μg/ml CTLA4-Ig are shown. ROT1D, recent-onset type 1 diabetes
    Ctla4 Ig, supplied by Bristol Myers, used in various techniques. Bioz Stars score: 92/100, based on 507 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Proteintech rabbit bax polyclonal
    Signature of therapeutic response in TN-09. ( a ) Analysis of the 1509 probe sets regulated between the <t>CTLA4-Ig</t> responders ( n = 8) and rapidly progressing placebo-treated individuals ( n = 7) matched for baseline I . I . 359 at the 3 month time point. The top three panels (left to right) illustrate two-way clustering of the selected and excluded participants in each arm. Left panel, mean expression of the four groups. Middle panel, participants selected for the analysis. Right panel, remaining CTLA4-Ig- and placebo-treated participants. Lower three panels, expression levels of a selected set of well-annotated transcripts. The colour bar indicates assigned treatment arm: red, placebo; blue, CTLA4-Ig. Note: samples were not available for two placebo-treated participants at the 3 month time point. The annotated dataset is available from the corresponding author on request. ( b ) Mean expression levels of the 1509 differentially induced probe sets at 12 and 24 months. ( c ) Upstream regulator analysis using IPA. A z score > 2.0 is significantly activated; a z score > −2.0 is significantly inhibited. ( d ) CTLA4-Ig add-back experiment. The mean induced expression levels of the 1509 probe sets after plasma of five local untreated individuals with diabetes were supplemented with 0 μg/ml, 25 μg/ml or 82 μg/ml CTLA4-Ig are shown. ROT1D, recent-onset type 1 diabetes
    Rabbit Bax Polyclonal, supplied by Proteintech, used in various techniques. Bioz Stars score: 99/100, based on 541 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Becton Dickinson ige
    H. polygyrus infection promotes a strong type 2 immune response. Real time qPCR analysis shows a clear suppression in the T-bet and RORγt mRNAs expression ( a and b ) and increase in the expression of GATA3 and Foxp3 mRNAs ( c and d ) in the MLN cells from HFD-fed H. polygyrus -infected mice. H. polygyrus -infected obese mice showed decreases in IFN-γ and IL-17 secretions in the supernatant of cultured MLN cells ( e and f ). H. polygyrus -infected obese mice had higher production of cytokines IL-4 and IL-10 in MLN ( g and h ). H. polygyrus -infected obese mice had increased levels of serum <t>IgE</t> ( i ) and <t>IgG1</t> ( j) and a decreased IgG2a level ( k ). (n = 5–6; * p
    Ige, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 94/100, based on 1480 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    TomoTherapy ig imrt
    Biochemical control rates between <t>HDR-BT</t> monotherapy and <t>IG-IMRT</t> with helical tomotherapy: ( a ) total population; ( b ) low-risk group; ( c ) intermediate-risk group; ( d ) high-risk group; ( e ) very-high-risk group.
    Ig Imrt, supplied by TomoTherapy, used in various techniques. Bioz Stars score: 90/100, based on 125 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    98
    Millipore anti dnp ige
    ( A ) Degranulation of SHIP+/+, +/−, and −/− BMMCs in response to the calcium ionophore A23187 or to <t>anti-DNP</t> <t>IgE</t> with or without DNP-HSA. Control values are those obtained with cells incubated for 15 min at 37°C in the absence of both IgE and DNP-HSA. The base 1-h IgE values are the controls for the IgE + DNP-HSA (in figure, see DNP). The black bars are the values obtained when 5 mM EGTA was added 1 min prior to IgE or DNP stimulation. Each bar is the mean of quadruplicates ±SD, and similar results were obtained in four separate experiments with three independently isolated lines of each genotype. ( B ) Intracellular Ca 2+ measurements in SHIP+/+, +/−, and −/− BMMCs in response to anti-DNP IgE + DNP-HSA with or without EGTA. ( C ) Intracellular Ca 2+ measurements in SHIP+/+, +/−, and −/− BMMCs in response to anti-DNP IgE alone with or without EGTA. The arrow indicates the time when DNP-HSA ( B ) or IgE ( C ) was added. The arrowhead depicts the time of EGTA (5 mM) addition. The Ca 2+ profiles of +/+ and +/− BMMCs obtained in the presence of EGTA (not shown) are similar to those shown with the −/− cells. Identical results were obtained in four separate experiments.
    Anti Dnp Ige, supplied by Millipore, used in various techniques. Bioz Stars score: 98/100, based on 380 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Thermo Fisher ige
    No cross-reactivity between bGST and HDM-GST. (A) IgG1-reactivity of sera (n = 3) collected from bGST-immunized mice at day 0 and day 59 to <t>BPE</t> and HDME. (B) <t>IgE-reactivity</t> of seven Der p 8-sensitized HDM-allergic patients to bGST and HDME. NHS, non-allergic control sera; O.D. optical density; dotted and dashed lines indicate the cut-off for positive IgE-reactivity.
    Ige, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 1875 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    SouthernBiotech goat anti mouse ig
    No cross-reactivity between bGST and HDM-GST. (A) IgG1-reactivity of sera (n = 3) collected from bGST-immunized mice at day 0 and day 59 to <t>BPE</t> and HDME. (B) <t>IgE-reactivity</t> of seven Der p 8-sensitized HDM-allergic patients to bGST and HDME. NHS, non-allergic control sera; O.D. optical density; dotted and dashed lines indicate the cut-off for positive IgE-reactivity.
    Goat Anti Mouse Ig, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 95/100, based on 308 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Proteintech mouse alpha tubulin monoclonal
    No cross-reactivity between bGST and HDM-GST. (A) IgG1-reactivity of sera (n = 3) collected from bGST-immunized mice at day 0 and day 59 to <t>BPE</t> and HDME. (B) <t>IgE-reactivity</t> of seven Der p 8-sensitized HDM-allergic patients to bGST and HDME. NHS, non-allergic control sera; O.D. optical density; dotted and dashed lines indicate the cut-off for positive IgE-reactivity.
    Mouse Alpha Tubulin Monoclonal, supplied by Proteintech, used in various techniques. Bioz Stars score: 99/100, based on 189 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    SouthernBiotech goat anti mouse
    No cross-reactivity between bGST and HDM-GST. (A) IgG1-reactivity of sera (n = 3) collected from bGST-immunized mice at day 0 and day 59 to <t>BPE</t> and HDME. (B) <t>IgE-reactivity</t> of seven Der p 8-sensitized HDM-allergic patients to bGST and HDME. NHS, non-allergic control sera; O.D. optical density; dotted and dashed lines indicate the cut-off for positive IgE-reactivity.
    Goat Anti Mouse, supplied by SouthernBiotech, used in various techniques. Bioz Stars score: 94/100, based on 508 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Millipore ige
    PMX-53 and CST do not activate murine peritoneal and bone marrow-derived mast cells. A, murine bone marrow-derived and peritoneal mast cells were incubated with <t>DNP-specific</t> mouse <t>IgE</t> (1 μg/ml, 16 h). Cells were exposed to buffer (control), 1
    Ige, supplied by Millipore, used in various techniques. Bioz Stars score: 96/100, based on 656 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Thermo Fisher total ige
    Serum levels of total <t>IgE</t> according to IL-21R gene polymorphisms in patients with KD. Our data showed no significant difference of serum total IgE between the patients who have IL-21R gene polymorphisms and those who have not (median value=30.1 pg/mL vs. 58.6 pg/mL, p=0.17). NS: not significant, IgE: immunoglobulin E, IL-21R: <t>interleukin-21</t> receptor, KD: Kawasaki disease.
    Total Ige, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 377 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Becton Dickinson mouse ige
    Serum levels of OVA-specific immunoglobulins. Serum samples of WT and mev/+ mice were collected within 2 hrs after challenge and OVA allergen-specific <t>IgE,</t> IgG1 and <t>IgG2a</t> were measured by ELISA. ( A ). Serum OVA-specific IgE. ( B ). Serum OVA-specific IgG1. ( C ). Serum OVA-specific IgG2a. (n = 5–7 mice each group; * P
    Mouse Ige, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 91/100, based on 218 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    Becton Dickinson rat anti mouse ige
    H . polygyrus/ P . chabaudi co-infection leads to impaired Th2 responses. A-C). Triple reporter mice were orally infected with 200 H . polygyrus larvae. 6 days post-infection, mice were infected i.p. with 10 5 P . chabaudi . At day 8 of P . chabaudi infection (d14 H . polygyrus ), mice were harvested. B). Total numbers of CD4 + CD44 hi Il4 gfp+ cells in the mesenteric lymph nodes. Data are representative of 5 independent experiments with 2–4 mice per group. C). <t>IgE</t> measured in the serum by <t>ELISA</t> from 3 pooled experiments. D and E). C57BL/6 mice were infected with 200 H . polygyrus larvae, treated on 2 consecutive days (days 14–15) with pyrantel pamoate (5 mg), infected with 10 5 P . chabaudi , and re-infected with H . polygyrus . Adult worms in intestine were counted on day 51. Data are representative of 4 independent experiments with 6–7 mice per group. * denotes P
    Rat Anti Mouse Ige, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 90/100, based on 247 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Vector Laboratories mouse ig blocking reagent
    H . polygyrus/ P . chabaudi co-infection leads to impaired Th2 responses. A-C). Triple reporter mice were orally infected with 200 H . polygyrus larvae. 6 days post-infection, mice were infected i.p. with 10 5 P . chabaudi . At day 8 of P . chabaudi infection (d14 H . polygyrus ), mice were harvested. B). Total numbers of CD4 + CD44 hi Il4 gfp+ cells in the mesenteric lymph nodes. Data are representative of 5 independent experiments with 2–4 mice per group. C). <t>IgE</t> measured in the serum by <t>ELISA</t> from 3 pooled experiments. D and E). C57BL/6 mice were infected with 200 H . polygyrus larvae, treated on 2 consecutive days (days 14–15) with pyrantel pamoate (5 mg), infected with 10 5 P . chabaudi , and re-infected with H . polygyrus . Adult worms in intestine were counted on day 51. Data are representative of 4 independent experiments with 6–7 mice per group. * denotes P
    Mouse Ig Blocking Reagent, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 99/100, based on 543 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Id1 is expressed in ECs and ST fibroblasts. a mRNA was isolated from HMVECs and fibroblasts were isolated from NL and RA ST. mRNA was reverse transcribed into cDNA and underwent PCR for 40 cycles. RA fibroblasts showed significantly elevated Id1 expression compared to NL ST fibroblasts and HMVECs. b Id1 is expressed in RA STs. IHC was performed on RA, OA, and NL ST cryosections. Tissues were blocked and then incubated using a mouse anti-human Id1 (Abcam) primary antibody. After washing, tissues were incubated with a biotinylated anti-mouse secondary antibody (Vector Laboratories). Tissues were washed and subsequently developed with the Vectastain ABC kit (Vector Laboratories). Id1 is found on synovial cells (SNC) in the RA ST. c For immunofluorescence staining, RA, osteoarthritis (OA), and normal (NL) ST sections were fixed in cold acetone for 30 min. The STs were blocked with 5 % donkey serum and 20 % fetal bovine serum (FBS) in PBS at 37 °C for 1 h, and then incubated with rabbit anti-human Id1 antibody (Abcam, 10 μg/ml) or purified nonspecific rabbit IgG for 1 h at 37 °C in blocking buffer. The synovial tissues samples were washed with PBS, and a 1:200 dilution in blocking buffer of fluorescent-conjugated donkey anti-rabbit antibody was added and incubated for an additional 1 h at 37 °C. As shown, we could validate Id1 staining in RA ST similar to what was found using IHC (×400). FLS fibroblast-like synoviocytes, HMVEC human dermal microvascular endothelial cell, Id1 inhibitor of DNA binding 1, IgG immunoglobulin, NL normal, RA rheumatoid arthritis, SNC synovial cell

    Journal: Arthritis Research & Therapy

    Article Title: Inflammatory properties of inhibitor of DNA binding 1 secreted by synovial fibroblasts in rheumatoid arthritis

    doi: 10.1186/s13075-016-0984-3

    Figure Lengend Snippet: Id1 is expressed in ECs and ST fibroblasts. a mRNA was isolated from HMVECs and fibroblasts were isolated from NL and RA ST. mRNA was reverse transcribed into cDNA and underwent PCR for 40 cycles. RA fibroblasts showed significantly elevated Id1 expression compared to NL ST fibroblasts and HMVECs. b Id1 is expressed in RA STs. IHC was performed on RA, OA, and NL ST cryosections. Tissues were blocked and then incubated using a mouse anti-human Id1 (Abcam) primary antibody. After washing, tissues were incubated with a biotinylated anti-mouse secondary antibody (Vector Laboratories). Tissues were washed and subsequently developed with the Vectastain ABC kit (Vector Laboratories). Id1 is found on synovial cells (SNC) in the RA ST. c For immunofluorescence staining, RA, osteoarthritis (OA), and normal (NL) ST sections were fixed in cold acetone for 30 min. The STs were blocked with 5 % donkey serum and 20 % fetal bovine serum (FBS) in PBS at 37 °C for 1 h, and then incubated with rabbit anti-human Id1 antibody (Abcam, 10 μg/ml) or purified nonspecific rabbit IgG for 1 h at 37 °C in blocking buffer. The synovial tissues samples were washed with PBS, and a 1:200 dilution in blocking buffer of fluorescent-conjugated donkey anti-rabbit antibody was added and incubated for an additional 1 h at 37 °C. As shown, we could validate Id1 staining in RA ST similar to what was found using IHC (×400). FLS fibroblast-like synoviocytes, HMVEC human dermal microvascular endothelial cell, Id1 inhibitor of DNA binding 1, IgG immunoglobulin, NL normal, RA rheumatoid arthritis, SNC synovial cell

    Article Snippet: After washing, tissues were incubated with a biotinylated anti-mouse or anti-rabbit secondary antibody (Vector Laboratories, Burlingame, CA, USA, 10 μg/mL) for 1 h at 37 °C in blocking buffer.

    Techniques: Isolation, Polymerase Chain Reaction, Expressing, Immunohistochemistry, Incubation, Plasmid Preparation, Immunofluorescence, Staining, Purification, Blocking Assay, Binding Assay

    SNC Id1 expression is significantly higher in inflamed ST and in the ankles of K/BxN serum-induced mice. a Id1 expression was visualized on SNCs in ST by IHC. A significantly greater percentage of SNCs were positive for Id1 in RA compared to OA and NL ST. b Percentages of cells expressing Id1 were also evaluated on SNCs from joint tissues taken from K/BxN serum-induced mice. A significantly greater percentage of SNCs positive for Id1 compared to Wt (noninduced mice) was found. c Murine tissues were blocked and then incubated using a rabbit anti-mouse Id1 antibody (CalBioreagents). After washing, tissues were incubated with a biotinylated anti-rabbit secondary antibody (Vector Laboratories). Tissues were washed and subsequently developed with the Vectastain ABC kit (Vector Laboratories). Id1 is found on SNCs near the bone in the K/BxN mouse ankles. Id1 inhibitor of DNA binding 1, IgG immunoglobulin, NL normal, OA osteoarthritis, RA rheumatoid arthritis, SNC synovial cell, ST synovial tissue, Wt wildtype

    Journal: Arthritis Research & Therapy

    Article Title: Inflammatory properties of inhibitor of DNA binding 1 secreted by synovial fibroblasts in rheumatoid arthritis

    doi: 10.1186/s13075-016-0984-3

    Figure Lengend Snippet: SNC Id1 expression is significantly higher in inflamed ST and in the ankles of K/BxN serum-induced mice. a Id1 expression was visualized on SNCs in ST by IHC. A significantly greater percentage of SNCs were positive for Id1 in RA compared to OA and NL ST. b Percentages of cells expressing Id1 were also evaluated on SNCs from joint tissues taken from K/BxN serum-induced mice. A significantly greater percentage of SNCs positive for Id1 compared to Wt (noninduced mice) was found. c Murine tissues were blocked and then incubated using a rabbit anti-mouse Id1 antibody (CalBioreagents). After washing, tissues were incubated with a biotinylated anti-rabbit secondary antibody (Vector Laboratories). Tissues were washed and subsequently developed with the Vectastain ABC kit (Vector Laboratories). Id1 is found on SNCs near the bone in the K/BxN mouse ankles. Id1 inhibitor of DNA binding 1, IgG immunoglobulin, NL normal, OA osteoarthritis, RA rheumatoid arthritis, SNC synovial cell, ST synovial tissue, Wt wildtype

    Article Snippet: After washing, tissues were incubated with a biotinylated anti-mouse or anti-rabbit secondary antibody (Vector Laboratories, Burlingame, CA, USA, 10 μg/mL) for 1 h at 37 °C in blocking buffer.

    Techniques: Expressing, Mouse Assay, Immunohistochemistry, Incubation, Plasmid Preparation, Binding Assay

    Formation of syncytia in CD4 + HeLa cells expressing CCR-5 and gp120/gp41 from a macrophage-tropic primary HIV-1 isolate. P4 (CCR-5 − ) ( 1 , 4 , 7 , and 8 ) or P4P (CCR-5 + ) ( 2 , 3 , 5 , and 6 ) HeLa cells (as indicated) were infected with defective recombinant SFV-LAI ( 1 , 2 , and 7 ), SFV-BX08 ( 4 , 5 , and 8 ) or SFV-LacZ ( 3 ). Control cells were left uninfected ( 6 ). After 12 h incubation at 37°C, cells were fixed with 0.5% (vol/vol) glutaraldehyde and stained with Giemsa ( 1 - 6 ) or reacted with mAb 2G12 ( 7 and 8 ) as primary antibody, followed by biotinylated anti-mouse IgG then avidin and biotinylated horseradish peroxidase H (Vectastain ABC kit). The cells were examined by microscopy. (×125.)

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Antibodies to several conformation-dependent epitopes of gp120/gp41 inhibit CCR-5-dependent cell-to-cell fusion mediated by the native envelope glycoprotein of a primary macrophage-tropic HIV-1 isolate

    doi:

    Figure Lengend Snippet: Formation of syncytia in CD4 + HeLa cells expressing CCR-5 and gp120/gp41 from a macrophage-tropic primary HIV-1 isolate. P4 (CCR-5 − ) ( 1 , 4 , 7 , and 8 ) or P4P (CCR-5 + ) ( 2 , 3 , 5 , and 6 ) HeLa cells (as indicated) were infected with defective recombinant SFV-LAI ( 1 , 2 , and 7 ), SFV-BX08 ( 4 , 5 , and 8 ) or SFV-LacZ ( 3 ). Control cells were left uninfected ( 6 ). After 12 h incubation at 37°C, cells were fixed with 0.5% (vol/vol) glutaraldehyde and stained with Giemsa ( 1 - 6 ) or reacted with mAb 2G12 ( 7 and 8 ) as primary antibody, followed by biotinylated anti-mouse IgG then avidin and biotinylated horseradish peroxidase H (Vectastain ABC kit). The cells were examined by microscopy. (×125.)

    Article Snippet: Virus stocks were titered on BHK cells, infection was detected by immunohistostaining with mAbs 41A and F5.5 (1 μg/ml) as primary antibodies, followed by biotinylated anti-mouse IgG then avidin and biotinylated horseradish peroxidase H (Vectastain ABC kit; Vector Laboratories).

    Techniques: Expressing, Infection, Recombinant, Incubation, Staining, Avidin-Biotin Assay, Microscopy

    Immunohistochemical (IHC) staining and hematoxylin and eosin (H E) staining After sacrifice of experimental mice, tumors were excised and fixed in 10% normal formalin. Samples were then stained with H E to confirm histopathological analysis and features of necrosis or apoptosis. Additionally, PCNA protein as a proliferation marker was detected in tissue specimens using a mouse monoclonal anti-PCNA primary antibody (1:100 dilution). Following incubation with primary antibody, biotinylated anti-mouse secondary antibody was applied to the slide (1:500 dilution). (A) Tumor mass of negative control (H E staining). (B) Tumor mass of CPT-11 treated mice (H E staining). (C) Tumor mass of HB1.F3.CE cells and CPT-11 co-treated mice (H E staining). (D) IHC staining for PCNA protein in each tumor mass. (E) The relative value of PCNA protein expression levels. *; p

    Journal: Oncotarget

    Article Title: Co-treatment with therapeutic neural stem cells expressing carboxyl esterase and CPT-11 inhibit growth of primary and metastatic lung cancers in mice

    doi:

    Figure Lengend Snippet: Immunohistochemical (IHC) staining and hematoxylin and eosin (H E) staining After sacrifice of experimental mice, tumors were excised and fixed in 10% normal formalin. Samples were then stained with H E to confirm histopathological analysis and features of necrosis or apoptosis. Additionally, PCNA protein as a proliferation marker was detected in tissue specimens using a mouse monoclonal anti-PCNA primary antibody (1:100 dilution). Following incubation with primary antibody, biotinylated anti-mouse secondary antibody was applied to the slide (1:500 dilution). (A) Tumor mass of negative control (H E staining). (B) Tumor mass of CPT-11 treated mice (H E staining). (C) Tumor mass of HB1.F3.CE cells and CPT-11 co-treated mice (H E staining). (D) IHC staining for PCNA protein in each tumor mass. (E) The relative value of PCNA protein expression levels. *; p

    Article Snippet: After four washes for 10 min in PBS-Tween, the slides were incubated in the biotinylated anti-mouse secondary antibody (1:500 dilution, Vector Laboratories) solution for 30 min at 37°C.

    Techniques: Immunohistochemistry, Staining, Mouse Assay, Marker, Incubation, Negative Control, Cycling Probe Technology, Expressing

    Photographic montage of Biotinylated Dextran Amine (BDA) injection sites into the DFL of PMv of lesion (cases 735, 830 and A42) and controls (1926, 1931 and 307A). The stimulation points from the ICMS surgery were coregistered with large blood vessels denoted by the asterisks on the immunohistochemically reacted tissue thereby verifying that the injection site was placed into the DFL of PMv. Each stimulation point was color-coded corresponding to a given body movement during ICMS (red, digits; green, wrist; blue, proximal arm movements; yellow, face, black, no response). Scale bars 2mm.

    Journal: bioRxiv

    Article Title: Anatomical Plasticity of the Distal Forelimb Projection of the Ventral Premotor Cortex Four weeks After Primary Motor Cortex Injury

    doi: 10.1101/2020.06.12.148494

    Figure Lengend Snippet: Photographic montage of Biotinylated Dextran Amine (BDA) injection sites into the DFL of PMv of lesion (cases 735, 830 and A42) and controls (1926, 1931 and 307A). The stimulation points from the ICMS surgery were coregistered with large blood vessels denoted by the asterisks on the immunohistochemically reacted tissue thereby verifying that the injection site was placed into the DFL of PMv. Each stimulation point was color-coded corresponding to a given body movement during ICMS (red, digits; green, wrist; blue, proximal arm movements; yellow, face, black, no response). Scale bars 2mm.

    Article Snippet: Following the 48 hour primary antibody incubation, tissue sections were rinsed and incubated at room temperature in a secondary biotinylated anti-mouse antibody for 4 hours (Vector Laboratories, Burlingame, CA).

    Techniques: Injection

    F-ATPase/VGCC protein complex verification. a. 1D immunoblot analysis of the VGCC α2δ-1 subunit in neutrophils. The VGCC α2δ-1 subunit-containing protein complexes in a NativePAGE Novex 4–16% Bis-Tris gel were detected at approximately 750 kDa via immunoblotting, the same position as the F-ATPase F1 β-subunit. b. 2D immunoblot analysis of the F-ATPase and VGCC protein complex. Plasma membrane lysed by 2% digitonin was loaded and run on a BN-PAGE gel. One lane (Sample) was cut and equilibrated in SDS loading buffer, followed by fixation on a SDS-PAGE gel for 2D electrophoretic analysis. Two separate immunoblot analyses with a primary antibody against the VGCC α2δ-1 subunit or the F-ATPase F1 β-subunit were performed. The F-ATPase F1 β-subunit appears vertically below the VGCC α2δ-1 subunit in the same PVDF membrane. c. Immunofluorescence colocalization analysis of F-ATPase and VGCC. Representative images show that the VGCC α2δ-1 subunit and F-ATPase F1 β-subunit colocalized on the plasma membrane of neutrophils. d. Immunoprecipitation of VGCC α2δ-1 subunit and F-ATPase from plasma membrane protein in human neutrophils. Samples were immunoprecipitated with a mouse monoclonal antibody against α2 of the VGCC α2δ-1 subunit. F-ATPase was detected with a mouse monoclonal antibody against the F-ATPase F1 α-subunit or β-subunit and an HRP-conjugated goat anti-mouse IgG light chain specifically bound antibody. IgG control experiment did not detect F-ATPase

    Journal: Cell Communication and Signaling : CCS

    Article Title: F0F1 ATP synthase regulates extracellular calcium influx in human neutrophils by interacting with Cav2.3 and modulates neutrophil accumulation in the lipopolysaccharide-challenged lung

    doi: 10.1186/s12964-020-0515-3

    Figure Lengend Snippet: F-ATPase/VGCC protein complex verification. a. 1D immunoblot analysis of the VGCC α2δ-1 subunit in neutrophils. The VGCC α2δ-1 subunit-containing protein complexes in a NativePAGE Novex 4–16% Bis-Tris gel were detected at approximately 750 kDa via immunoblotting, the same position as the F-ATPase F1 β-subunit. b. 2D immunoblot analysis of the F-ATPase and VGCC protein complex. Plasma membrane lysed by 2% digitonin was loaded and run on a BN-PAGE gel. One lane (Sample) was cut and equilibrated in SDS loading buffer, followed by fixation on a SDS-PAGE gel for 2D electrophoretic analysis. Two separate immunoblot analyses with a primary antibody against the VGCC α2δ-1 subunit or the F-ATPase F1 β-subunit were performed. The F-ATPase F1 β-subunit appears vertically below the VGCC α2δ-1 subunit in the same PVDF membrane. c. Immunofluorescence colocalization analysis of F-ATPase and VGCC. Representative images show that the VGCC α2δ-1 subunit and F-ATPase F1 β-subunit colocalized on the plasma membrane of neutrophils. d. Immunoprecipitation of VGCC α2δ-1 subunit and F-ATPase from plasma membrane protein in human neutrophils. Samples were immunoprecipitated with a mouse monoclonal antibody against α2 of the VGCC α2δ-1 subunit. F-ATPase was detected with a mouse monoclonal antibody against the F-ATPase F1 α-subunit or β-subunit and an HRP-conjugated goat anti-mouse IgG light chain specifically bound antibody. IgG control experiment did not detect F-ATPase

    Article Snippet: Voltage-gated calcium channel (VGCC) α2δ-1 was detected with a rabbit polyclonal antibody against the VGCC α2δ-1 subunit (1:200, C5105, Sigma-Aldrich, St. Louis, MO, USA) and an HRP-conjugated goat anti-rabbit IgG H & L (1:2000, ab6721, Abcam, Cambridge, UK).

    Techniques: Polyacrylamide Gel Electrophoresis, SDS Page, Immunofluorescence, Immunoprecipitation

    miR-137 represses cholangiocarcinoma cell proliferation in vitro . (A) Fluorescence microscope examination (magnification, ×200) and reverse transcription-quantitative PCR were used to detect the infection efficiency of miR-137 overexpression (LV-miR-137) and NC lentiviruses. (B) The effect of miR-137 on cholangiocarcinoma cell proliferation was detected by the Cell Counting Kit-8 assay. (C) The effect of miR-137 on cholangiocarcinoma cell colony formation ability was detected using colony formation assays. (D) Cell cycle distribution was analyzed after miR-137 overexpression in TFK-1 and HuCCT1 cells. (E) Western blotting was used to detect the expression of CDK2 and cyclin D1 in the miR-137 overexpression and normal control groups. GAPDH was used as the loading control. * P

    Journal: International Journal of Molecular Medicine

    Article Title: MicroRNA-137 suppresses the proliferation, migration and invasion of cholangiocarcinoma cells by targeting WNT2B

    doi: 10.3892/ijmm.2020.4474

    Figure Lengend Snippet: miR-137 represses cholangiocarcinoma cell proliferation in vitro . (A) Fluorescence microscope examination (magnification, ×200) and reverse transcription-quantitative PCR were used to detect the infection efficiency of miR-137 overexpression (LV-miR-137) and NC lentiviruses. (B) The effect of miR-137 on cholangiocarcinoma cell proliferation was detected by the Cell Counting Kit-8 assay. (C) The effect of miR-137 on cholangiocarcinoma cell colony formation ability was detected using colony formation assays. (D) Cell cycle distribution was analyzed after miR-137 overexpression in TFK-1 and HuCCT1 cells. (E) Western blotting was used to detect the expression of CDK2 and cyclin D1 in the miR-137 overexpression and normal control groups. GAPDH was used as the loading control. * P

    Article Snippet: After blocking with TBST (0.1% Tween-20) containing 5% skimmed milk at room temperature for 2 h, the membranes were then incubated with primary antibodies (1:1,000) against WNT2B (cat. no. ab50575; Abcam), β-catenin (cat. no. 51067-2-AP; Proteintech), TCF4 (cat. no. 22337-1-AP; Proteintech), N-cadherin (cat. no. 22018-1-AP; Proteintech), vimentin (cat. no. 10366-1-AP; Proteintech), cyclin D1 (cat. no. 26939-1-AP; Proteintech), CDK2 (cat. no. 10122-1-AP; Proteintech), c-Myc (cat. no. 10828-1-AP; Proteintech) and GAPDH (cat. no. 60004-1-Ig; Proteintech) for 16 h at 4°C.

    Techniques: In Vitro, Fluorescence, Microscopy, Real-time Polymerase Chain Reaction, Infection, Over Expression, Cell Counting, Western Blot, Expressing

    TDP-43 OE can reduce the mRNA and protein levels of Parkin in an intron-independent manner. a – d TDP-43 selectively reduces the mRNA levels of plasmid-expressed, intron-free Parkin but not PINK1 . The mRNA levels of transiently transfected Flag-Parkin ( a ) or PINK1-V5 ( c ) in 293T cells are evaluated by a semiquantitative RT-PCR assay, and quantified in b and d , respectively. The specific PCR primers (containing the Flag or V5 tag sequence) are designed to distinguish the plasmid mRNAs from endogenously expressed Parkin and PINK1 mRNAs in 293T cells. e , f TDP-43 OE decreases the protein levels of Parkin but increases cleaved PINK1. Western blot analysis of protein levels of plasmid-expressed Flag-Parkin ( e ) or PINK1-V5 ( g ) in 293T cells, and quantified in f and h , respectively. All protein levels are normalized to GAPDH. en-TDP-43 endogenous human TDP-43 in 293T cells, tr-TDP-43 transfected hTDP-43-HA, f-PINK1 full-length PINK1, c-PINK1 cleaved PINK1. Data are means ± SEM; n = 3–5; * p

    Journal: Cell Death & Disease

    Article Title: Distinct multilevel misregulations of Parkin and PINK1 revealed in cell and animal models of TDP-43 proteinopathy

    doi: 10.1038/s41419-018-1022-y

    Figure Lengend Snippet: TDP-43 OE can reduce the mRNA and protein levels of Parkin in an intron-independent manner. a – d TDP-43 selectively reduces the mRNA levels of plasmid-expressed, intron-free Parkin but not PINK1 . The mRNA levels of transiently transfected Flag-Parkin ( a ) or PINK1-V5 ( c ) in 293T cells are evaluated by a semiquantitative RT-PCR assay, and quantified in b and d , respectively. The specific PCR primers (containing the Flag or V5 tag sequence) are designed to distinguish the plasmid mRNAs from endogenously expressed Parkin and PINK1 mRNAs in 293T cells. e , f TDP-43 OE decreases the protein levels of Parkin but increases cleaved PINK1. Western blot analysis of protein levels of plasmid-expressed Flag-Parkin ( e ) or PINK1-V5 ( g ) in 293T cells, and quantified in f and h , respectively. All protein levels are normalized to GAPDH. en-TDP-43 endogenous human TDP-43 in 293T cells, tr-TDP-43 transfected hTDP-43-HA, f-PINK1 full-length PINK1, c-PINK1 cleaved PINK1. Data are means ± SEM; n = 3–5; * p

    Article Snippet: The following antibodies were used for western blotting, immunoprecipitation, and immunofluorescence assays: mouse anti-FLAG (Sigma-Aldrich, F3165); mouse anti-HA (Proteintech, 66006-1); rabbit anti-HA (CST, 3724); rabbit anti-TDP-43 (Proteintech, 10782-2-AP); rabbit anti-PINK1 (Novus Biologicals, BC100-494); mouse anti-Parkin (Santa Cruz, sc-32282); mouse anti-V5 (Proteintech, 66007-1); mouse anti-GAPDH (Proteintech, 60004-1); rabbit anti-Tubulin (MBL, PM054); mouse anti-Lamin C (DSHB, LC28.26); and chicken anti-MAP2 (Abcam, ab5392).

    Techniques: Plasmid Preparation, Transfection, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Sequencing, Western Blot

    RNP purification from animal tissue. Mouse brain tissue was cryo-grinded and UV-irradiated before (Hot)-PTex was performed. Western blot against HuR (ELAVL1) demonstrates recovery of cross-linked HuR from mouse tissue while beta-actin (ACTB) is efficiently depleted. For full blots see Supplementary Figure 15

    Journal: Nature Communications

    Article Title: Purification of cross-linked RNA-protein complexes by phenol-toluol extraction

    doi: 10.1038/s41467-019-08942-3

    Figure Lengend Snippet: RNP purification from animal tissue. Mouse brain tissue was cryo-grinded and UV-irradiated before (Hot)-PTex was performed. Western blot against HuR (ELAVL1) demonstrates recovery of cross-linked HuR from mouse tissue while beta-actin (ACTB) is efficiently depleted. For full blots see Supplementary Figure 15

    Article Snippet: Primary antibodies targeted the proteins HuR (1:1000, Proteintech, 11910–1-AP), ABCF2 (1:1000, Proteintech, 10226-1-AP), CCT7 (1:1000, Proteintech, 15994-1-AP), FUS (1:1000, abcam, ab124923), GAPDH (1:1000, Proteintech, 10494-1-AP), alpha-enolase (ENO1, 1:1000, Proteintech, 11204-1-AP), PTBP1 (1:1000, abcam, ab133734), PABPC1 (1:1000, Proteintech, 10970-1-AP), ACTB (1:1000, Proteintech, 66009-1-Ig), Histone H3 (1:1000, abcam, ab21054), FLAG-tag (1:5000, Sigma, A8592), or Sxl-RBD4 (DHSB, anti-Sxl hybridome culture supernatant M114, 1:20).

    Techniques: Purification, Irradiation, Western Blot

    Signature of therapeutic response in TN-09. ( a ) Analysis of the 1509 probe sets regulated between the CTLA4-Ig responders ( n = 8) and rapidly progressing placebo-treated individuals ( n = 7) matched for baseline I . I . 359 at the 3 month time point. The top three panels (left to right) illustrate two-way clustering of the selected and excluded participants in each arm. Left panel, mean expression of the four groups. Middle panel, participants selected for the analysis. Right panel, remaining CTLA4-Ig- and placebo-treated participants. Lower three panels, expression levels of a selected set of well-annotated transcripts. The colour bar indicates assigned treatment arm: red, placebo; blue, CTLA4-Ig. Note: samples were not available for two placebo-treated participants at the 3 month time point. The annotated dataset is available from the corresponding author on request. ( b ) Mean expression levels of the 1509 differentially induced probe sets at 12 and 24 months. ( c ) Upstream regulator analysis using IPA. A z score > 2.0 is significantly activated; a z score > −2.0 is significantly inhibited. ( d ) CTLA4-Ig add-back experiment. The mean induced expression levels of the 1509 probe sets after plasma of five local untreated individuals with diabetes were supplemented with 0 μg/ml, 25 μg/ml or 82 μg/ml CTLA4-Ig are shown. ROT1D, recent-onset type 1 diabetes

    Journal: Diabetologia

    Article Title: Innate immune activity as a predictor of persistent insulin secretion and association with responsiveness to CTLA4-Ig treatment in recent-onset type 1 diabetes

    doi: 10.1007/s00125-018-4708-x

    Figure Lengend Snippet: Signature of therapeutic response in TN-09. ( a ) Analysis of the 1509 probe sets regulated between the CTLA4-Ig responders ( n = 8) and rapidly progressing placebo-treated individuals ( n = 7) matched for baseline I . I . 359 at the 3 month time point. The top three panels (left to right) illustrate two-way clustering of the selected and excluded participants in each arm. Left panel, mean expression of the four groups. Middle panel, participants selected for the analysis. Right panel, remaining CTLA4-Ig- and placebo-treated participants. Lower three panels, expression levels of a selected set of well-annotated transcripts. The colour bar indicates assigned treatment arm: red, placebo; blue, CTLA4-Ig. Note: samples were not available for two placebo-treated participants at the 3 month time point. The annotated dataset is available from the corresponding author on request. ( b ) Mean expression levels of the 1509 differentially induced probe sets at 12 and 24 months. ( c ) Upstream regulator analysis using IPA. A z score > 2.0 is significantly activated; a z score > −2.0 is significantly inhibited. ( d ) CTLA4-Ig add-back experiment. The mean induced expression levels of the 1509 probe sets after plasma of five local untreated individuals with diabetes were supplemented with 0 μg/ml, 25 μg/ml or 82 μg/ml CTLA4-Ig are shown. ROT1D, recent-onset type 1 diabetes

    Article Snippet: Briefly, induced transcription for each participant at 3, 12 and 24 months was normalised with that of baseline, then differentially induced transcripts between the CTLA4-Ig and placebo arms were identified.

    Techniques: Expressing, Indirect Immunoperoxidase Assay

    Type 1 diabetes subgroups exhibit different rates of C-peptide decline and responses to CTLA4-Ig treatment. A significant inverse relationship between baseline I . I . 359 and the rate (slope) of C-peptide decline in placebo-treated TN-09 participants is described in Fig. 2 f. Here, this relationship is considered from the perspective of subgroups 1–4. The green data point is the outlying CTLA4-Ig participant not assigned to a subgroup (left-most participant in Fig. 6 d). Regression lines are shown for the CTLA4-Ig and placebo arms of TN-09 in blue and red, respectively

    Journal: Diabetologia

    Article Title: Innate immune activity as a predictor of persistent insulin secretion and association with responsiveness to CTLA4-Ig treatment in recent-onset type 1 diabetes

    doi: 10.1007/s00125-018-4708-x

    Figure Lengend Snippet: Type 1 diabetes subgroups exhibit different rates of C-peptide decline and responses to CTLA4-Ig treatment. A significant inverse relationship between baseline I . I . 359 and the rate (slope) of C-peptide decline in placebo-treated TN-09 participants is described in Fig. 2 f. Here, this relationship is considered from the perspective of subgroups 1–4. The green data point is the outlying CTLA4-Ig participant not assigned to a subgroup (left-most participant in Fig. 6 d). Regression lines are shown for the CTLA4-Ig and placebo arms of TN-09 in blue and red, respectively

    Article Snippet: Briefly, induced transcription for each participant at 3, 12 and 24 months was normalised with that of baseline, then differentially induced transcripts between the CTLA4-Ig and placebo arms were identified.

    Techniques:

    Type 1 diabetes subgroups exhibit different responses to CTLA4-Ig treatment. ( a ) Heatmaps illustrating expression levels of the 916 significantly regulated transcripts belonging to the three modules significantly correlated with 2 h C-peptide AUC at baseline and the placebo-treated arm of TN-09; an annotated heatmap is shown below. The four major subgroups (as shown in Fig. 6 d) are indicated for each arm. Well-annotated transcripts are illustrated, and the module to which they belong is indicated by the colour bar on the left. Colour bars are provided that indicate: fold change; baseline C-peptide AUC; rate of C-peptide decline over 24 months; selected CTLA4-Ig responders (as in Fig. 5 ; blue); selected placebos (as in Fig. 5 ; ‘Selected fast placebos’; peach); age at baseline; and I . I . 359 . ( b ) TN-09 participants belonging to subgroups 1–4 exhibit different responses to CTLA4-Ig treatment. Tabulated are subgroup mean values for each metric. Statistical differences were assessed with a two-tailed unpaired Student’s t test, superscript letters denote p

    Journal: Diabetologia

    Article Title: Innate immune activity as a predictor of persistent insulin secretion and association with responsiveness to CTLA4-Ig treatment in recent-onset type 1 diabetes

    doi: 10.1007/s00125-018-4708-x

    Figure Lengend Snippet: Type 1 diabetes subgroups exhibit different responses to CTLA4-Ig treatment. ( a ) Heatmaps illustrating expression levels of the 916 significantly regulated transcripts belonging to the three modules significantly correlated with 2 h C-peptide AUC at baseline and the placebo-treated arm of TN-09; an annotated heatmap is shown below. The four major subgroups (as shown in Fig. 6 d) are indicated for each arm. Well-annotated transcripts are illustrated, and the module to which they belong is indicated by the colour bar on the left. Colour bars are provided that indicate: fold change; baseline C-peptide AUC; rate of C-peptide decline over 24 months; selected CTLA4-Ig responders (as in Fig. 5 ; blue); selected placebos (as in Fig. 5 ; ‘Selected fast placebos’; peach); age at baseline; and I . I . 359 . ( b ) TN-09 participants belonging to subgroups 1–4 exhibit different responses to CTLA4-Ig treatment. Tabulated are subgroup mean values for each metric. Statistical differences were assessed with a two-tailed unpaired Student’s t test, superscript letters denote p

    Article Snippet: Briefly, induced transcription for each participant at 3, 12 and 24 months was normalised with that of baseline, then differentially induced transcripts between the CTLA4-Ig and placebo arms were identified.

    Techniques: Expressing, Two Tailed Test

    The relationship between I . I . 359 and age of clinical onset. ( a ) Among the 74 TN-09 participants analysed, the age at diagnosis was directly related to baseline 2 h C-peptide AUC ( r 2 = 0.28, p = 2.1 × 10 −5 ). The baseline I . I . 359 was independent of age in TN-09 ( b ) and local ( c ) participants ( n = 26). In ( a , b ), black solid circles, placebo treated ( n = 19); grey open circles, CTLA4-Ig treated ( n = 54); dotted/dashed vertical line, 18 years of age

    Journal: Diabetologia

    Article Title: Innate immune activity as a predictor of persistent insulin secretion and association with responsiveness to CTLA4-Ig treatment in recent-onset type 1 diabetes

    doi: 10.1007/s00125-018-4708-x

    Figure Lengend Snippet: The relationship between I . I . 359 and age of clinical onset. ( a ) Among the 74 TN-09 participants analysed, the age at diagnosis was directly related to baseline 2 h C-peptide AUC ( r 2 = 0.28, p = 2.1 × 10 −5 ). The baseline I . I . 359 was independent of age in TN-09 ( b ) and local ( c ) participants ( n = 26). In ( a , b ), black solid circles, placebo treated ( n = 19); grey open circles, CTLA4-Ig treated ( n = 54); dotted/dashed vertical line, 18 years of age

    Article Snippet: Briefly, induced transcription for each participant at 3, 12 and 24 months was normalised with that of baseline, then differentially induced transcripts between the CTLA4-Ig and placebo arms were identified.

    Techniques:

    Th17 cells express higher levels of CTLA-4 and are less susceptible to CTLA-4 Ig than Th1 cells Draining popliteal LNs from M.Tb OVA and Candida OVA mice were collected on day 9 post-immunization and (A–D) restimulated for 4 h with PMA/Iono or (E–F) co-cultured with OVA 323–339 and DCs in the presence of CTLA-4 Ig or IgG-Fc for 4 d before brief PMA/Iono restimulation. Data shown depict Th1 and Th7 OT-II cells from both immunization groups (n = 16–21/group, 3–4 experiments). (A) The frequency of CD28 + and (B) CD28 expression level on antigen-specific Th1 and Th17 cells (p = n.s.). (C) The frequency of CTLA-4 + (p = 0.0006) and (D) CTLA-4 expression level (p = 0.0044) on OT-II Th1 and Th17 cells. (E) CD11c + DCs were purified from naïve splenocytes and analyzed for MHC Class II I-A b , CD80, and CD86 expression. (F) M.Tb OVA and Candida OVA CD4 + T cells were co-cultured with DCs for 4 d in the presence of CTLA-4 Ig IgG-Fc control followed by brief PMA/Iono restimulation. (F) Representative frequencies (top number) and absolute numbers (bottom number) of OT-II Th1 and Th17 cells without PMA/Iono restimulation (first row) or with restimulation (middle/bottom rows, p = 0.0015). Bar graphs include average ± SEM. Statistical comparison performed using (A–D) unpaired or (F) paired two-tailed Student’s t-test. **p

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: Candida-Elicited Murine Th17 Cells Express High CTLA-4 Compared to Th1 Cells and Are Resistant to Costimulation Blockade

    doi: 10.4049/jimmunol.1301332

    Figure Lengend Snippet: Th17 cells express higher levels of CTLA-4 and are less susceptible to CTLA-4 Ig than Th1 cells Draining popliteal LNs from M.Tb OVA and Candida OVA mice were collected on day 9 post-immunization and (A–D) restimulated for 4 h with PMA/Iono or (E–F) co-cultured with OVA 323–339 and DCs in the presence of CTLA-4 Ig or IgG-Fc for 4 d before brief PMA/Iono restimulation. Data shown depict Th1 and Th7 OT-II cells from both immunization groups (n = 16–21/group, 3–4 experiments). (A) The frequency of CD28 + and (B) CD28 expression level on antigen-specific Th1 and Th17 cells (p = n.s.). (C) The frequency of CTLA-4 + (p = 0.0006) and (D) CTLA-4 expression level (p = 0.0044) on OT-II Th1 and Th17 cells. (E) CD11c + DCs were purified from naïve splenocytes and analyzed for MHC Class II I-A b , CD80, and CD86 expression. (F) M.Tb OVA and Candida OVA CD4 + T cells were co-cultured with DCs for 4 d in the presence of CTLA-4 Ig IgG-Fc control followed by brief PMA/Iono restimulation. (F) Representative frequencies (top number) and absolute numbers (bottom number) of OT-II Th1 and Th17 cells without PMA/Iono restimulation (first row) or with restimulation (middle/bottom rows, p = 0.0015). Bar graphs include average ± SEM. Statistical comparison performed using (A–D) unpaired or (F) paired two-tailed Student’s t-test. **p

    Article Snippet: After 4 days, the absolute number of Th1 cells was significantly diminished by CTLA-4 Ig compared to Ig control (37.7±5.4% of Ig-Fc, ).

    Techniques: Mouse Assay, Cell Culture, Expressing, Purification, Two Tailed Test

    Candida polarized mice have greater neutrophil recruitment to skin grafts in the presence of CTLA-4 Ig M.Tb OVA and Candida OVA mice received an OVA skin transplant on day 14 post-immunization. Mice were left untreated or were treated with 500 µg each CTLA-4 Ig and anti-CD154. On day 6 post-graft skin grafts were harvested and snap frozen for histological analysis of (A) macrophage or (B) neutrophil infiltration. Cellular infiltration was quantified using Aperio ImageScope software (n = 20–22 image fields/group). Bar graphs show average±SEM. Statistical analysis was performed using one-way ANOVA with Bonferroni post test. *p

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: Candida-Elicited Murine Th17 Cells Express High CTLA-4 Compared to Th1 Cells and Are Resistant to Costimulation Blockade

    doi: 10.4049/jimmunol.1301332

    Figure Lengend Snippet: Candida polarized mice have greater neutrophil recruitment to skin grafts in the presence of CTLA-4 Ig M.Tb OVA and Candida OVA mice received an OVA skin transplant on day 14 post-immunization. Mice were left untreated or were treated with 500 µg each CTLA-4 Ig and anti-CD154. On day 6 post-graft skin grafts were harvested and snap frozen for histological analysis of (A) macrophage or (B) neutrophil infiltration. Cellular infiltration was quantified using Aperio ImageScope software (n = 20–22 image fields/group). Bar graphs show average±SEM. Statistical analysis was performed using one-way ANOVA with Bonferroni post test. *p

    Article Snippet: After 4 days, the absolute number of Th1 cells was significantly diminished by CTLA-4 Ig compared to Ig control (37.7±5.4% of Ig-Fc, ).

    Techniques: Mouse Assay, Software

    Th17 Cells are more susceptible to CTLA-4 coinhibition than Th1 cells Draining popliteal LNs from M.Tb OVA and Candida OVA mice were collected on day 9 post-immunization and co-cultured with OVA 323–339 and DCs in the presence of anti-CTLA-4 or IgG for 4 d before brief PMA/Iono restimulation. Data shown depict OT-II Th1 and Th7 cells from both immunization groups (n = 13–15/group, 3 experiments). (A) Representative frequencies (top number) and absolute numbers (bottom number) of OT-II Th1 and Th17 cells without PMA/Iono restimulation (first row) or with restimulation (middle/bottom rows) (B) Th1 and Th17 cell numbers from M.Tb OVA and Candida OVA groups following anti-CTLA-4 blockade (p = 0.0387). Bar graphs depict average ± SEM. Statistical comparison performed using unpaired two-tailed Student’s t-test *p

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: Candida-Elicited Murine Th17 Cells Express High CTLA-4 Compared to Th1 Cells and Are Resistant to Costimulation Blockade

    doi: 10.4049/jimmunol.1301332

    Figure Lengend Snippet: Th17 Cells are more susceptible to CTLA-4 coinhibition than Th1 cells Draining popliteal LNs from M.Tb OVA and Candida OVA mice were collected on day 9 post-immunization and co-cultured with OVA 323–339 and DCs in the presence of anti-CTLA-4 or IgG for 4 d before brief PMA/Iono restimulation. Data shown depict OT-II Th1 and Th7 cells from both immunization groups (n = 13–15/group, 3 experiments). (A) Representative frequencies (top number) and absolute numbers (bottom number) of OT-II Th1 and Th17 cells without PMA/Iono restimulation (first row) or with restimulation (middle/bottom rows) (B) Th1 and Th17 cell numbers from M.Tb OVA and Candida OVA groups following anti-CTLA-4 blockade (p = 0.0387). Bar graphs depict average ± SEM. Statistical comparison performed using unpaired two-tailed Student’s t-test *p

    Article Snippet: After 4 days, the absolute number of Th1 cells was significantly diminished by CTLA-4 Ig compared to Ig control (37.7±5.4% of Ig-Fc, ).

    Techniques: Mouse Assay, Cell Culture, Two Tailed Test

    Candida elicited CD4 + T cells can mediate costimulation blockade resistant graft rejection (A) Mice were adoptively transferred with 10 6 OT-I and OT-II cells 24 h before immunization with M.Tb or Candida and OVA peptide. (B–C) Draining popliteal LNs were isolated on (B) day 9 or (C) day 14 post-immunization and the frequency and absolute number of of Thy1.1 + OT-II cells were determined (p = n.s., n = 10–12/group, 2 experiments). (D–E) At 14 d post-immunization, mice were transplanted with mOVA skin and monitored for graft survival (n = 13–14/group, 3 experiments). (D) Mice were left untreated (p = n.s.) or (E) treated with 500 µg each CTLA-4 Ig and anti-CD154 (p = 0.0002). Statistical comparisons performed using unpaired two-tailed Student’s t-test (B) and log rank test (D–E), *p

    Journal: Journal of immunology (Baltimore, Md. : 1950)

    Article Title: Candida-Elicited Murine Th17 Cells Express High CTLA-4 Compared to Th1 Cells and Are Resistant to Costimulation Blockade

    doi: 10.4049/jimmunol.1301332

    Figure Lengend Snippet: Candida elicited CD4 + T cells can mediate costimulation blockade resistant graft rejection (A) Mice were adoptively transferred with 10 6 OT-I and OT-II cells 24 h before immunization with M.Tb or Candida and OVA peptide. (B–C) Draining popliteal LNs were isolated on (B) day 9 or (C) day 14 post-immunization and the frequency and absolute number of of Thy1.1 + OT-II cells were determined (p = n.s., n = 10–12/group, 2 experiments). (D–E) At 14 d post-immunization, mice were transplanted with mOVA skin and monitored for graft survival (n = 13–14/group, 3 experiments). (D) Mice were left untreated (p = n.s.) or (E) treated with 500 µg each CTLA-4 Ig and anti-CD154 (p = 0.0002). Statistical comparisons performed using unpaired two-tailed Student’s t-test (B) and log rank test (D–E), *p

    Article Snippet: After 4 days, the absolute number of Th1 cells was significantly diminished by CTLA-4 Ig compared to Ig control (37.7±5.4% of Ig-Fc, ).

    Techniques: Mouse Assay, Isolation, Two Tailed Test

    H. polygyrus infection promotes a strong type 2 immune response. Real time qPCR analysis shows a clear suppression in the T-bet and RORγt mRNAs expression ( a and b ) and increase in the expression of GATA3 and Foxp3 mRNAs ( c and d ) in the MLN cells from HFD-fed H. polygyrus -infected mice. H. polygyrus -infected obese mice showed decreases in IFN-γ and IL-17 secretions in the supernatant of cultured MLN cells ( e and f ). H. polygyrus -infected obese mice had higher production of cytokines IL-4 and IL-10 in MLN ( g and h ). H. polygyrus -infected obese mice had increased levels of serum IgE ( i ) and IgG1 ( j) and a decreased IgG2a level ( k ). (n = 5–6; * p

    Journal: Scientific Reports

    Article Title: Helminth infection protects against high fat diet-induced obesity via induction of alternatively activated macrophages

    doi: 10.1038/s41598-018-22920-7

    Figure Lengend Snippet: H. polygyrus infection promotes a strong type 2 immune response. Real time qPCR analysis shows a clear suppression in the T-bet and RORγt mRNAs expression ( a and b ) and increase in the expression of GATA3 and Foxp3 mRNAs ( c and d ) in the MLN cells from HFD-fed H. polygyrus -infected mice. H. polygyrus -infected obese mice showed decreases in IFN-γ and IL-17 secretions in the supernatant of cultured MLN cells ( e and f ). H. polygyrus -infected obese mice had higher production of cytokines IL-4 and IL-10 in MLN ( g and h ). H. polygyrus -infected obese mice had increased levels of serum IgE ( i ) and IgG1 ( j) and a decreased IgG2a level ( k ). (n = 5–6; * p

    Article Snippet: Measurement of serum antibody Each mouse was bled at sacrifice, and individual sera were assayed for total IgE, IgG1 and IgG2a by ELISA on plates coated with goat anti-mouse Ab to IgG1, IgG2a (BD PharMingen) or rat anti-mouse Ab to IgE (BD PharMingen).

    Techniques: Infection, Real-time Polymerase Chain Reaction, Expressing, Mouse Assay, Cell Culture

    Biochemical control rates between HDR-BT monotherapy and IG-IMRT with helical tomotherapy: ( a ) total population; ( b ) low-risk group; ( c ) intermediate-risk group; ( d ) high-risk group; ( e ) very-high-risk group.

    Journal: Cancers

    Article Title: High-Dose-Rate Brachytherapy Monotherapy versus Image-Guided Intensity-Modulated Radiotherapy with Helical Tomotherapy for Patients with Localized Prostate Cancer

    doi: 10.3390/cancers10090322

    Figure Lengend Snippet: Biochemical control rates between HDR-BT monotherapy and IG-IMRT with helical tomotherapy: ( a ) total population; ( b ) low-risk group; ( c ) intermediate-risk group; ( d ) high-risk group; ( e ) very-high-risk group.

    Article Snippet: The actuarial five-year biochemical failure-free survival rate was 92.9% (95% confidential interval (CI) = 90.1–95.6%) and 89.2% (95% CI = 85.9–92.9%, p = 0.18; p = 0.077 in IPTW) for HDR-BT and IG-IMRT, respectively; they were 97.3% (100% for HDR-BT and 95.8% for IG-IMRT, p = 0.28; p = 0.15 in IPTW) for the low-risk group, 94.2% (95.6% for HDR-BT and 92% for IG-IMRT, p = 0.42; p = 0.60 in IPTW) for the intermediate-risk group, 87.7% (90.4% for HDR-BT and 84.9% for IG-IMRT, p = 0.10; p = 0.041 in IPTW) for the high-risk group, and 87.8% (87.1% for HDR-BT and 89.2% for IG-IMRT, p = 0.38; p = 0.60 in IPTW) for the very-high-risk group.

    Techniques:

    ( A ) Degranulation of SHIP+/+, +/−, and −/− BMMCs in response to the calcium ionophore A23187 or to anti-DNP IgE with or without DNP-HSA. Control values are those obtained with cells incubated for 15 min at 37°C in the absence of both IgE and DNP-HSA. The base 1-h IgE values are the controls for the IgE + DNP-HSA (in figure, see DNP). The black bars are the values obtained when 5 mM EGTA was added 1 min prior to IgE or DNP stimulation. Each bar is the mean of quadruplicates ±SD, and similar results were obtained in four separate experiments with three independently isolated lines of each genotype. ( B ) Intracellular Ca 2+ measurements in SHIP+/+, +/−, and −/− BMMCs in response to anti-DNP IgE + DNP-HSA with or without EGTA. ( C ) Intracellular Ca 2+ measurements in SHIP+/+, +/−, and −/− BMMCs in response to anti-DNP IgE alone with or without EGTA. The arrow indicates the time when DNP-HSA ( B ) or IgE ( C ) was added. The arrowhead depicts the time of EGTA (5 mM) addition. The Ca 2+ profiles of +/+ and +/− BMMCs obtained in the presence of EGTA (not shown) are similar to those shown with the −/− cells. Identical results were obtained in four separate experiments.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: The src homology 2-containing inositol phosphatase (SHIP) is the gatekeeper of mast cell degranulation

    doi:

    Figure Lengend Snippet: ( A ) Degranulation of SHIP+/+, +/−, and −/− BMMCs in response to the calcium ionophore A23187 or to anti-DNP IgE with or without DNP-HSA. Control values are those obtained with cells incubated for 15 min at 37°C in the absence of both IgE and DNP-HSA. The base 1-h IgE values are the controls for the IgE + DNP-HSA (in figure, see DNP). The black bars are the values obtained when 5 mM EGTA was added 1 min prior to IgE or DNP stimulation. Each bar is the mean of quadruplicates ±SD, and similar results were obtained in four separate experiments with three independently isolated lines of each genotype. ( B ) Intracellular Ca 2+ measurements in SHIP+/+, +/−, and −/− BMMCs in response to anti-DNP IgE + DNP-HSA with or without EGTA. ( C ) Intracellular Ca 2+ measurements in SHIP+/+, +/−, and −/− BMMCs in response to anti-DNP IgE alone with or without EGTA. The arrow indicates the time when DNP-HSA ( B ) or IgE ( C ) was added. The arrowhead depicts the time of EGTA (5 mM) addition. The Ca 2+ profiles of +/+ and +/− BMMCs obtained in the presence of EGTA (not shown) are similar to those shown with the −/− cells. Identical results were obtained in four separate experiments.

    Article Snippet: If stimulating with IgE + DNP, cells were incubated at 4°C with anti-DNP IgE for 1 h, washed twice with 23°C Tyrode’s buffer, equilibrated in Tyrode’s buffer to 37°C for 5 min, and then treated for 15 min with or without (base 1 h IgE) DNP-human serum albumin (DNP-HSA; Sigma).

    Techniques: Incubation, Isolation

    Tyrosine phosphorylations following IgE sensitization. SHIP+/+, +/−, and −/− BMMCs were starved for 2–4 h and then incubated either at 37°C for 3 min with 10 μg/ml anti-DNP IgE or at 4°C for 1 h with 10 μg/ml IgE followed by two washes and a 3 min incubation with 20 ng/ml DNP-HSA at 37°C. Cells were then washed, solubilized, and subjected to immunoprecipitation (IP) with anti-FcɛR1 β-subunit ( A ) , anti-SHIP ( B; ) , or anti-Shc antibodies ( C ). The blots were reprobed with the immunoprecipitating antibody to show equal loading. ( D ) Total cell lysates from SHIP+/+ and −/− BMMCs with or without EGTA were incubated for 1 min before the 3-min stimulation with IgE or DNP-HSA and then subjected to Western analysis with anti-phospho-MAPK (New England BioLabs) and reprobed with anti-ERK 1-CT (Upstate Biotechnology) to confirm equal loading. The last three lanes of both the +/+ and −/− samples are total cell lysates from cells treated for 1 h with IgE at 4°C, shifted up to 37°C and treated with or without EGTA for 1 min before stimulating for 3 min with DNP-HSA.

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: The src homology 2-containing inositol phosphatase (SHIP) is the gatekeeper of mast cell degranulation

    doi:

    Figure Lengend Snippet: Tyrosine phosphorylations following IgE sensitization. SHIP+/+, +/−, and −/− BMMCs were starved for 2–4 h and then incubated either at 37°C for 3 min with 10 μg/ml anti-DNP IgE or at 4°C for 1 h with 10 μg/ml IgE followed by two washes and a 3 min incubation with 20 ng/ml DNP-HSA at 37°C. Cells were then washed, solubilized, and subjected to immunoprecipitation (IP) with anti-FcɛR1 β-subunit ( A ) , anti-SHIP ( B; ) , or anti-Shc antibodies ( C ). The blots were reprobed with the immunoprecipitating antibody to show equal loading. ( D ) Total cell lysates from SHIP+/+ and −/− BMMCs with or without EGTA were incubated for 1 min before the 3-min stimulation with IgE or DNP-HSA and then subjected to Western analysis with anti-phospho-MAPK (New England BioLabs) and reprobed with anti-ERK 1-CT (Upstate Biotechnology) to confirm equal loading. The last three lanes of both the +/+ and −/− samples are total cell lysates from cells treated for 1 h with IgE at 4°C, shifted up to 37°C and treated with or without EGTA for 1 min before stimulating for 3 min with DNP-HSA.

    Article Snippet: If stimulating with IgE + DNP, cells were incubated at 4°C with anti-DNP IgE for 1 h, washed twice with 23°C Tyrode’s buffer, equilibrated in Tyrode’s buffer to 37°C for 5 min, and then treated for 15 min with or without (base 1 h IgE) DNP-human serum albumin (DNP-HSA; Sigma).

    Techniques: Incubation, Immunoprecipitation, Western Blot

    No cross-reactivity between bGST and HDM-GST. (A) IgG1-reactivity of sera (n = 3) collected from bGST-immunized mice at day 0 and day 59 to BPE and HDME. (B) IgE-reactivity of seven Der p 8-sensitized HDM-allergic patients to bGST and HDME. NHS, non-allergic control sera; O.D. optical density; dotted and dashed lines indicate the cut-off for positive IgE-reactivity.

    Journal: PLoS ONE

    Article Title: Glutathione-S-Transferase: A Minor Allergen in Birch Pollen due to Limited Release from Hydrated Pollen

    doi: 10.1371/journal.pone.0109075

    Figure Lengend Snippet: No cross-reactivity between bGST and HDM-GST. (A) IgG1-reactivity of sera (n = 3) collected from bGST-immunized mice at day 0 and day 59 to BPE and HDME. (B) IgE-reactivity of seven Der p 8-sensitized HDM-allergic patients to bGST and HDME. NHS, non-allergic control sera; O.D. optical density; dotted and dashed lines indicate the cut-off for positive IgE-reactivity.

    Article Snippet: Briefly, all patients suffered from rhinoconjunctivitis in spring and showed positive skin prick test responses to BPE and specific IgE to BP (≥0.35 kUA /L, ImmunoCAP, Thermofisher, Uppsala, Sweden).

    Techniques: Mouse Assay

    IgE-responses of BP-allergic individuals. (A) IgE-reactivity of 217 BP-allergic patients to BP allergens and bGST. (B) IgE-reactivity to bGST is inhibited by pre-incubation of sera with BPE (gray line) and bGST (black line). One representative example of four is shown. (C) RBL assay with sera from three patients sensitized to bGST and Bet v 1. Degranulation was induced with titrated amounts of bGST (filled circles) and Bet v 1 (open circles). The release of β-hexosaminidase is expressed as percentage of the release from anti-IgE-treated cells.

    Journal: PLoS ONE

    Article Title: Glutathione-S-Transferase: A Minor Allergen in Birch Pollen due to Limited Release from Hydrated Pollen

    doi: 10.1371/journal.pone.0109075

    Figure Lengend Snippet: IgE-responses of BP-allergic individuals. (A) IgE-reactivity of 217 BP-allergic patients to BP allergens and bGST. (B) IgE-reactivity to bGST is inhibited by pre-incubation of sera with BPE (gray line) and bGST (black line). One representative example of four is shown. (C) RBL assay with sera from three patients sensitized to bGST and Bet v 1. Degranulation was induced with titrated amounts of bGST (filled circles) and Bet v 1 (open circles). The release of β-hexosaminidase is expressed as percentage of the release from anti-IgE-treated cells.

    Article Snippet: Briefly, all patients suffered from rhinoconjunctivitis in spring and showed positive skin prick test responses to BPE and specific IgE to BP (≥0.35 kUA /L, ImmunoCAP, Thermofisher, Uppsala, Sweden).

    Techniques: Incubation

    PMX-53 and CST do not activate murine peritoneal and bone marrow-derived mast cells. A, murine bone marrow-derived and peritoneal mast cells were incubated with DNP-specific mouse IgE (1 μg/ml, 16 h). Cells were exposed to buffer (control), 1

    Journal: Molecular Pharmacology

    Article Title: PMX-53 as a Dual CD88 Antagonist and an Agonist for Mas-Related Gene 2 (MrgX2) in Human Mast Cells

    doi: 10.1124/mol.111.071472

    Figure Lengend Snippet: PMX-53 and CST do not activate murine peritoneal and bone marrow-derived mast cells. A, murine bone marrow-derived and peritoneal mast cells were incubated with DNP-specific mouse IgE (1 μg/ml, 16 h). Cells were exposed to buffer (control), 1

    Article Snippet: Monoclonal anti-DNP specific IgE and anti-human IgE were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Derivative Assay, Incubation

    PMX-53 uses MrgX2 to induce mast cell degranulation. RBL-2H3 cells stably expressing MrgX1 (A) or MrgX2 (B) were incubated with anti-DNP specific IgE (1 μg/ml, 16 h) and stimulated with the indicated peptides (1 μM), substance P (SP; 1

    Journal: Molecular Pharmacology

    Article Title: PMX-53 as a Dual CD88 Antagonist and an Agonist for Mas-Related Gene 2 (MrgX2) in Human Mast Cells

    doi: 10.1124/mol.111.071472

    Figure Lengend Snippet: PMX-53 uses MrgX2 to induce mast cell degranulation. RBL-2H3 cells stably expressing MrgX1 (A) or MrgX2 (B) were incubated with anti-DNP specific IgE (1 μg/ml, 16 h) and stimulated with the indicated peptides (1 μM), substance P (SP; 1

    Article Snippet: Monoclonal anti-DNP specific IgE and anti-human IgE were purchased from Sigma-Aldrich (St. Louis, MO).

    Techniques: Stable Transfection, Expressing, Incubation

    Serum levels of total IgE according to IL-21R gene polymorphisms in patients with KD. Our data showed no significant difference of serum total IgE between the patients who have IL-21R gene polymorphisms and those who have not (median value=30.1 pg/mL vs. 58.6 pg/mL, p=0.17). NS: not significant, IgE: immunoglobulin E, IL-21R: interleukin-21 receptor, KD: Kawasaki disease.

    Journal: Korean Circulation Journal

    Article Title: Interleukin-21 Receptor Gene Polymorphisms in Kawasaki Disease

    doi: 10.4070/kcj.2013.43.1.38

    Figure Lengend Snippet: Serum levels of total IgE according to IL-21R gene polymorphisms in patients with KD. Our data showed no significant difference of serum total IgE between the patients who have IL-21R gene polymorphisms and those who have not (median value=30.1 pg/mL vs. 58.6 pg/mL, p=0.17). NS: not significant, IgE: immunoglobulin E, IL-21R: interleukin-21 receptor, KD: Kawasaki disease.

    Article Snippet: Measurement of interleukin-21 and total IgE in sera We measured the serum IL-21 level in patients with KD by enzyme linked immunosorbent assay kits (e-Bioscience, San Diego, CA, USA) and total IgE by immunoassay system (Siemens, Munich, Germany).

    Techniques:

    Serum levels of OVA-specific immunoglobulins. Serum samples of WT and mev/+ mice were collected within 2 hrs after challenge and OVA allergen-specific IgE, IgG1 and IgG2a were measured by ELISA. ( A ). Serum OVA-specific IgE. ( B ). Serum OVA-specific IgG1. ( C ). Serum OVA-specific IgG2a. (n = 5–7 mice each group; * P

    Journal: PLoS ONE

    Article Title: SHP-1 Regulation of Mast Cell Function in Allergic Inflammation and Anaphylaxis

    doi: 10.1371/journal.pone.0055763

    Figure Lengend Snippet: Serum levels of OVA-specific immunoglobulins. Serum samples of WT and mev/+ mice were collected within 2 hrs after challenge and OVA allergen-specific IgE, IgG1 and IgG2a were measured by ELISA. ( A ). Serum OVA-specific IgE. ( B ). Serum OVA-specific IgG1. ( C ). Serum OVA-specific IgG2a. (n = 5–7 mice each group; * P

    Article Snippet: Purified mouse IgE and IgG2a were used as standards (BD Pharmingen) and the levels of IgE and IgG2a were calculated accordingly.

    Techniques: Mouse Assay, Enzyme-linked Immunosorbent Assay

    H . polygyrus/ P . chabaudi co-infection leads to impaired Th2 responses. A-C). Triple reporter mice were orally infected with 200 H . polygyrus larvae. 6 days post-infection, mice were infected i.p. with 10 5 P . chabaudi . At day 8 of P . chabaudi infection (d14 H . polygyrus ), mice were harvested. B). Total numbers of CD4 + CD44 hi Il4 gfp+ cells in the mesenteric lymph nodes. Data are representative of 5 independent experiments with 2–4 mice per group. C). IgE measured in the serum by ELISA from 3 pooled experiments. D and E). C57BL/6 mice were infected with 200 H . polygyrus larvae, treated on 2 consecutive days (days 14–15) with pyrantel pamoate (5 mg), infected with 10 5 P . chabaudi , and re-infected with H . polygyrus . Adult worms in intestine were counted on day 51. Data are representative of 4 independent experiments with 6–7 mice per group. * denotes P

    Journal: PLoS Pathogens

    Article Title: IFNγ and IL-12 Restrict Th2 Responses during Helminth/Plasmodium Co-Infection and Promote IFNγ from Th2 Cells

    doi: 10.1371/journal.ppat.1004994

    Figure Lengend Snippet: H . polygyrus/ P . chabaudi co-infection leads to impaired Th2 responses. A-C). Triple reporter mice were orally infected with 200 H . polygyrus larvae. 6 days post-infection, mice were infected i.p. with 10 5 P . chabaudi . At day 8 of P . chabaudi infection (d14 H . polygyrus ), mice were harvested. B). Total numbers of CD4 + CD44 hi Il4 gfp+ cells in the mesenteric lymph nodes. Data are representative of 5 independent experiments with 2–4 mice per group. C). IgE measured in the serum by ELISA from 3 pooled experiments. D and E). C57BL/6 mice were infected with 200 H . polygyrus larvae, treated on 2 consecutive days (days 14–15) with pyrantel pamoate (5 mg), infected with 10 5 P . chabaudi , and re-infected with H . polygyrus . Adult worms in intestine were counted on day 51. Data are representative of 4 independent experiments with 6–7 mice per group. * denotes P

    Article Snippet: Total IgE ELISA was performed by coating with Purified Rat Anti-Mouse IgE (R35-72, BD Pharmingen) at 2 μg/mL overnight, followed by overnight incubation with serum and standard (Purified Mouse IgE,k isotype Standard, BD Pharmingen), and detection with Biotin Rat Anti-Mouse IgE at 1 μg/mL (R35-118, BD Pharmingen), Streptavidin HRP at 1:000 (BD Pharmingen) and ABTS One Component HRP Microwell Substrate (SurModics).

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

    Blockade of IL-12 and IFNγ during co-infection preserves Th2 responses. A-C). C57BL/6 mice were orally infected with 200 H . polygyrus larvae. 6 days post-infection, mice were infected with 10 5 P . chabaudi . At day 8-post infection with P . chabaudi (d14 H . polygyrus ), mice were harvested. Mice were treated with 0.5 mg of anti-IL-12 and anti-IFNγ i.p. at days 0, 5, and 11. B). Total numbers of CD4 + CD44 hi Il4 gfp+ cells in the mesenteric lymph nodes. C). IgE measured in the serum by ELISA. Data are representative of 2 independent experiments with 6 mice per group. * denotes P

    Journal: PLoS Pathogens

    Article Title: IFNγ and IL-12 Restrict Th2 Responses during Helminth/Plasmodium Co-Infection and Promote IFNγ from Th2 Cells

    doi: 10.1371/journal.ppat.1004994

    Figure Lengend Snippet: Blockade of IL-12 and IFNγ during co-infection preserves Th2 responses. A-C). C57BL/6 mice were orally infected with 200 H . polygyrus larvae. 6 days post-infection, mice were infected with 10 5 P . chabaudi . At day 8-post infection with P . chabaudi (d14 H . polygyrus ), mice were harvested. Mice were treated with 0.5 mg of anti-IL-12 and anti-IFNγ i.p. at days 0, 5, and 11. B). Total numbers of CD4 + CD44 hi Il4 gfp+ cells in the mesenteric lymph nodes. C). IgE measured in the serum by ELISA. Data are representative of 2 independent experiments with 6 mice per group. * denotes P

    Article Snippet: Total IgE ELISA was performed by coating with Purified Rat Anti-Mouse IgE (R35-72, BD Pharmingen) at 2 μg/mL overnight, followed by overnight incubation with serum and standard (Purified Mouse IgE,k isotype Standard, BD Pharmingen), and detection with Biotin Rat Anti-Mouse IgE at 1 μg/mL (R35-118, BD Pharmingen), Streptavidin HRP at 1:000 (BD Pharmingen) and ABTS One Component HRP Microwell Substrate (SurModics).

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