Journal: Nature

Article Title: Local immune response to food antigens drives meal-induced abdominal pain

doi: 10.1038/s41586-020-03118-2

Figure Lengend Snippet: a, Experimental protocol. b, Diarrhea score in OVA/sham + OVA and OVA/infected + OVA mice (n = 6 and 8, respectively). c, Quantification of OVA-specific IgE in serum (left) and colon homogenates (right) of OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline, OVA/infected + OVA and OVA-allergy mice (serum: n = 9, 10, 10, 9 and 8, respectively; colon: n = 9, 6, 9, 9 and 7, respectively). d-h, VMR to colorectal distention in (d) OVA/sham + OVA, OVA/infected + saline, saline/infected + OVA and OVA/infected + OVA mice (n = 12, 11, 11 and 13, respectively); (e) mice OVA-tolerized (high-dose) + saline/infected + OVA, OVA-tolerized (high-dose) + OVA/infected + OVA, OVA-tolerized (low-dose) + saline/infected + OVA and OVA-tolerized (low-dose) + OVA/infected + OVA (n = 6, 9, 6 and 8, respectively); (f) OVA/infected + OVA mice treated with anti-IgE antibody or control antibody (n = 8/group); (g); OVA/infected + OVA mice with WT or Igh7-/- background (n = 10/group); (h) and naïve mice treated with monoclonal OVA-specific IgE antibody or monoclonal Dinitrophenyl (DNP) antibody (n = 7 and 6, respectively). i, mIL-4-forming cells in coMLN from saline/infected + OVA, OVA/infected + OVA and OVA/infected + OVA mice (n = 8/group). Two-tailed Mann-Whitney test in b; Kruskal-Wallis test (Dunn’s multiple-comparisons test) in c and i; two-way repeated ANOVA (Sidak’s multiple-comparisons test) in d-h. Data shown as violin plots in b and median ± IQR in c-i. VMR, visceromotor response; BL, baseline; w, week.

Article Snippet: Monoclonal OVA-specific IgE antibody Mouse anti-OVA monoclonal IgE antibody (clone E-C1, Chondrex) was diluted in PBS.

Techniques: Infection, Two Tailed Test, MANN-WHITNEY

Journal: Nature

Article Title: Local immune response to food antigens drives meal-induced abdominal pain

doi: 10.1038/s41586-020-03118-2

Figure Lengend Snippet: a, b, diarrhea development quantification by (a) water content in feces and (b) whole-gut transit time upon gavage of carmine red dye in OVA/sham + OVA, OVA/infected + OVA (n = 10/group) mice. c, quantification of OVA-specific IgE in intestinal homogenates of OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA mice (n = 9, 10, 10 and 9, respectively) at 7 weeks post-infection. d, ear-swelling after intradermal injection of OVA in OVA/sham + OVA (n = 6), OVA/infected + OVA (n = 10), at 7 weeks post-infection, and OVA-allergy mice (n = 6, 10 and 6, respectively). e, f, VMR to colorectal distention in (e) OVA/sham + OVA, OVA/infected + saline, saline/infected + OVA and OVA/infected + OVA mice (n = 12, 11, 11 and 13, respectively) and (f) tracing of a electromyographic response to 20-μL-, 40-μL-, 60-μL- and 80-μL-volume colorectal distention in a OVA/infected + OVA mouse at baseline and 7 weeks post-infection. g, VMR to colorectal distention in OVA/infected + OVA (n = 7) and OVA/infected = Saline (n = 7 and 5, respectively) mice at baseline (BL), 7 weeks post-infection (PI) and after 1, 2, 3 and 4 weeks after stopping oral OVA or saline re-exposure, respectively. h, colonic permeability in BALB/c mice expressed as passage of fluorescein sodium (left) and transepithelial resistance (right) in OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA mice (n = 8, 11, 12 and 8, respectively) at 7 weeks post-infection. i, scheme illustrating the post-infectious protocol with prior OVA tolerization. j, VMR to colorectal distention in mice OVA-tolerized (high-dose) + saline/infected + OVA, OVA-tolerized (high-dose) + OVA/infected + OVA, OVA-tolerized (low-dose) + saline/infected + OVA and OVA-tolerized (low-dose) + OVA/infected + OVA (n = 6, 9, 6 and 8, respectively). k, l, VMR to colorectal distention in mice OVA/infected repeatedly gavaged with BSA compared to OVA (n = 6 and 10, respectively). m, scheme illustrating the post-infectious protocol in mice treated with anti-IgE antibody. n, VMR to colorectal distention in OVA/infected + OVA mice treated with anti-IgE antibody or control antibody (n = 8/group). o, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial resistance (right) of (l) VHS mice treated with anti-IgE antibody or control antibody (n = 8/group). p, VMR to colorectal distention in OVA/infected + OVA mice with WT or Igh7-/- background (n = 10/group). q, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial resistance (right) of OVA/infected + OVA mice with Igh7-/- background or WT mice (n = 10/group) at 7 weeks post-infection. r, scheme illustrating the protocol in mice that received monoclonal OVA-specific IgE antibody. s, VMR to colorectal distention in naïve mice treated with monoclonal OVA-specific IgE antibody or monoclonal Dinitrophenyl (DNP) antibody (n = 7 and 6, respectively). Two-tailed Mann–Whitney test in a and b for every time point, and in o and q (left); two-tailed t-test for q (right). Kruskal-Wallis test (Dunn’s multiple-comparisons test) in c and d. One-way ANOVA (Sidak’s multiple-comparisons test) in h. Two-way repeated ANOVA (Sidak’s multiple-comparisons test) in e, g, j-l, n, p and s. Data shown as Box-and-whiskers (center line, median; box, 25th-75th percentiles; whiskers, 10th-90th percentiles) in a, b and d and median ± IQR in c, e, g, h, j-l, n-q and s. VMR, visceromotor response; BL, baseline; w, week; d; day.

Article Snippet: Monoclonal OVA-specific IgE antibody Mouse anti-OVA monoclonal IgE antibody (clone E-C1, Chondrex) was diluted in PBS.

Techniques: Infection, Injection, Permeability, Two Tailed Test, MANN-WHITNEY

Journal: Nature

Article Title: Local immune response to food antigens drives meal-induced abdominal pain

doi: 10.1038/s41586-020-03118-2

Figure Lengend Snippet: Colonic compliance (volume-pressure relationship) evaluated in animals subjected to visceromotor response recordings. ns = non-significant.

Article Snippet: Monoclonal OVA-specific IgE antibody Mouse anti-OVA monoclonal IgE antibody (clone E-C1, Chondrex) was diluted in PBS.

Techniques:

Journal: Nature

Article Title: Local immune response to food antigens drives meal-induced abdominal pain

doi: 10.1038/s41586-020-03118-2

Figure Lengend Snippet: a-c, avidin-fluorescence intensity over time (expressed as TCCF) in MC from OVA/sham + OVA, saline/infected + OVA OVA/infected + saline and OVA/infected + OVA mice (n = 222 [7 mice], 145 [6 mice], 202 [8 mice] and 239 [9 mice], respectively), (a) shown as percentage at times = 0, 7, 14, 21 and 28 min, (b) micrographs and (c) shown at time = 28 min as median per mouse. In b, arrows point to representative MC showing degranulation (yellow, 28 min) compared to baseline (white, 0 min). d, representative image of double-staining using DAPI and avidin in fixed colonic tissue from healthy mice (n = 3). e, Histamine quantification in supernatant collected from OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA (n = 10/group) at 7 weeks post-infection. f, VMR to colorectal distention in OVA/infected + OVA mice (d) treated with doxantrazole or vehicle (n = 14 and 11, respectively). g, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of OVA/infected + OVA mice treated with doxantrazole or vehicle (n = 8/group) at 8 weeks post-infection. h, quantification of population of CD117+FcRεI+ mast cells in intestinal lamina propria from Cpa3Cre/+ and WT mice (n = 3/group). i, VMR to colorectal distention in OVA/infected + OVA mice with Cpa3Cre/+ background or WT littermates (n = 13 and 12, respectively). j, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) in OVA/infected + OVA mice with Cpa3Cre /+ background or WT littermates (n = 12/group) at 9 weeks post-infection. k, scheme illustrating the post-infectious protocol in mice treated with anti-CD20 antibody and bortezomib. l, anti-CD20 (5D2 clone) antibody and bortezomib depleted CD19+ Ly6K+ plasma cells (left) and B220+ IgM+ B cells (right) from the colon of mice compared to control antibody/vehicle and naïve mice (n = 2, 1 and 4, respectively) and (m) gating strategy used to validate B cell and plasma cell depletion. n, quantification of OVA-specific IgE in colon samples homogenates from OVA/infected + OVA mice treated with anti-CD20 antibody and bortezomib or control antibody and vehicle (n = 12 and 9, respectively). o, VMR to colorectal distention in OVA/infected + OVA mice treated with anti-CD20 antibody/bortezomib or control antibody/vehicle (n = 3 and 4, respectively). Mixed-effects model (Dunnett’s multiple-comparisons test) in a and e. One-way ANOVA (Sidak’s multiple-comparisons test) in c. two-way repeated measures ANOVA (Sidak’s multiple-comparisons test) in f, I and o. Two-tailed Mann-Whitney in g, h, j and n. Data are shown as median ± IQR in a, e-j, l, n and o, and box-and-whiskers (center line, median; box, 25th and 75th percentiles; whiskers, 10th and 90th percentiles) in c. VMR, visceromotor response; BL, baseline; w, week.

Article Snippet: Monoclonal OVA-specific IgE antibody Mouse anti-OVA monoclonal IgE antibody (clone E-C1, Chondrex) was diluted in PBS.

Techniques: Activation Assay, Avidin-Biotin Assay, Fluorescence, Infection, Double Staining, Permeability, Two Tailed Test, MANN-WHITNEY

Journal: Nature

Article Title: Local immune response to food antigens drives meal-induced abdominal pain

doi: 10.1038/s41586-020-03118-2

Figure Lengend Snippet: a, scheme illustrating the SEB protocol. b, VMR to colorectal distention in saline/SEB + OVA, OVA/SEB + OVA and OVA (low dose) + OVA/SEB + OVA mice (n = 10, 14 and 4, respectively). c, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of saline/SEB + OVA and OVA/SEB + OVA (n = 8 and 7, respectively) mice. d, heat-map of the gene expression of inflammatory genes and mast-cell-related genes in OVA/saline + OVA (a), saline/SEB + OVA (b) and OVA/SEB + OVA mice (c) (n = 21, 7 and 7, respectively; except for Tryptase α/β-1: n = 7/group). e, VMR to colorectal distention in SEB-VHS mice treated with doxantrazole or vehicle (n = 8 and 9, respectively). f, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of SEB-VHS mice treated with doxantrazole or vehicle (n = 8 and 7, respectively). g, VMR to colorectal distention in OVA/SEB + OVA mice with Cpa3Cre/+ background or WT littermates (n = 5 and 6, respectively). h, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of OVA/SEB + OVA mice with Cpa3Cre/+ background or WT littermates (n = 5 and 6, respectively). i, quantification of OVA-specific IgE in colon homogenates of saline/SEB + OVA and OVA/SEB + OVA mice (n = 8 and 7, respectively). j, ear-swelling after intradermal injection of OVA in saline/SEB + OVA, OVA/SEB + OVA and OVA-allergy mice (n = 9, 7 and 6, respectively) mice. k, % of HV and IBS patients (n = 64 and 84, respectively) positive for S. aureus (left) and SAg-encoding S. aureus (right) in fecal samples. Numbers below the bars represent the ratio of positive and total HV and patients. Two-way repeated ANOVA (Sidak’s multiple-comparisons test) in b, e and g. Two-tailed unpaired t-test in c and f. Two-tailed Mann-Whitney test in h and i. Kruskal-Wallis test (Dunn’s multiple-comparisons test) in d and j. Two-sided Fisher’s exact test in k. Data shown as median ± IQR in b, e-i and box-and-whiskers (center line, median; box, 25th-75th percentiles; whiskers, 10th-90th percentiles) in j. VMR, visceromotor response; BL, baseline; w, week. In d, Tpsab1, Il4, Il6 and Il10 gene expression differences were statistically significant in OVA/SEB + OVA vs. OVA/saline + OVA (adjusted p = 0.0005, 0.0004, 0.0163 and 0.0039, respectively).

Article Snippet: Monoclonal OVA-specific IgE antibody Mouse anti-OVA monoclonal IgE antibody (clone E-C1, Chondrex) was diluted in PBS.

Techniques: Infection, Permeability, Expressing, Injection, Two Tailed Test, MANN-WHITNEY

Journal: Nature

Article Title: Local immune response to food antigens drives meal-induced abdominal pain

doi: 10.1038/s41586-020-03118-2

Figure Lengend Snippet: Bacterial infection (or bacterial toxins, SEB) can trigger break of oral tolerance to food antigens leading to food-induced VHS upon food-antigen re-exposure. OVA-specific IgE antibodies bind to and sensitizes tissue-resident mast cells, which are activated upon re-exposure to OVA during feeding and release mediators that sensitize afferent neurons via H1R-mediated pathway.

Article Snippet: Monoclonal OVA-specific IgE antibody Mouse anti-OVA monoclonal IgE antibody (clone E-C1, Chondrex) was diluted in PBS.

Techniques: Infection

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: Dose dependency of each IgE monoclonal antibody against CPE in a sandwich ELISA.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques: Sandwich ELISA

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: Dose dependency of each IgE monoclonal antibody against CPE in an indirect ELISA.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques: Indirect ELISA

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: Immuno-blot analysis of anti-CPE IgE monoclonal antibodies against CPE. A. SDS-PAGE and Immuno-blot analysis of CPE under non-reducing conditions (Left), reducing conditions (Right). Coomassie brilliant blue R-250 staining; M: Molecular Marker and 1: CPE.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques: SDS Page, Staining, Marker

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: RBL-2H3 cell degranulation with CPE and anti-CPE IgE monoclonal antibodies.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques:

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: Dose dependency of CPE in RBL-2H3 cells degranulation with Anti-CPE monoclonal antibodies. RBL-2H3 cells were cultured in 0.1 ml of DMEM containing 15 % FBS at 10^6 cells/well in a 96-well plate for 3 hours, and then treated with anti-CPE IgE mAbs: 6E10C12 (Circle), 2G11G7 (Triangle), 6B7B10 (Square), and 5E7B12 (Diamond) at 37°C for 16 hours. After washing cells with PBS two times, various concentrations of CPE in Tyrode's buffer were added at 200 µl/well and then incubated at 37°C for 1 hour. 0.1 ml of medium from each well was transferred to a 96-well plate and assayed for beta-hexosaminidase activity. The degranulation of RBL-2H3 cells was expressed as a ratio compared with 100 % degranulation of the cells which received 1 % Triton-X in Tyrode's buffer. Data is expressed as the mean ± standard deviation.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques: Cell Culture, Incubation, Activity Assay, Standard Deviation

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: Footpad type I hypersensitivity by anti-CPE IgE monoclonal antibodies.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques:

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: Dose dependency of footpad type I hypersensitivity by monoclonal antibody, clone 2G11G7. The footpad thickness of mice after IV administration of each dose of mAbs: 0.1 mg (Circle), 0.3 mg (Square), and 1 mg (Triangle), followed by a 0.1 mg CPE (solid line) or PBS (dashed line) ID administration at the footpad. Data is expressed as the mean ± standard deviation.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques: Standard Deviation

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet: Summary of the Application of Anti-CPE IgE Monoclonal Antibodies.

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques: Indirect ELISA, Sandwich ELISA

Journal: MethodsX

Article Title: Development and evaluation of mouse anti-Ara h 1 and Ara h 3 IgE monoclonal antibodies for advancing peanut allergy research

doi: 10.1016/j.mex.2023.102470

Figure Lengend Snippet:

Article Snippet: After washing the plates with PBST, biotin-labeled anti-mouse IgE monoclonal antibodies (Chondrex) diluted in 0.05M Tris-buffered saline buffer, pH 8.0 containing 2 % Casein (Sigma-Aldrich, Inc., MO, USA) and 0.05 % Tween 20 (Casein/TBS) were added to the plates and incubated for 1 hour at RT.

Techniques:

Journal: Korean Journal of Pediatrics

Article Title: Hu.4-1BB-Fc fusion protein inhibits allergic inflammation and airway hyperresponsiveness in a murine model of asthma

doi: 10.3345/kjp.2011.54.9.373

Figure Lengend Snippet: Levels of total immunoglobulin (Ig) E, ovalbumin (OVA)-specific IgE, IgG 1 , and IgG 2a as determined by enzyme-linked immunosorbent assay in the sera of each group ( * P <0.05 and † P <0.01)

Article Snippet: Antibodies for OVA-specific IgE and total IgE (Acris antibodies GmbH, Herford, Germany) and IgG 1 and IgG 2a (Santa Cruz Biotechnology, Santa Cruz, CA, USA) were used.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Bioscience Reports

Article Title: The final proteolytic step in transmembrane signaling of multiple RsgI anti-σ factors in Clostridium thermocellum

doi: 10.1042/BSR20253055

Figure Lengend Snippet: (A ) SDS-PAGE of the eight purified RsgI-NFs samples. The expected RsgI-NF bands are indicated by red arrows. ( B ) Western blot analysis of the eight purified RsgI-NFs using the anti-His6-tag antibodies.

Article Snippet: These membranes were blocked using TBST-blocking buffer (50 mM Tris, 150 mM NaCl, 0.07% Tween-20, 5% skim milk, pH 7.4) for a duration of 2 h. Then, the membranes were incubated with mouse anti-His tag monoclonal antibody (Bioss Antibodies, Beijing, China) overnight at 4°C.

Techniques: SDS Page, Purification, Western Blot