rabbit polyclonal anti-mouse muc2  (Santa Cruz Biotechnology)

Journal: Cell reports

Article Title: Dynamic genetic adaptation of Bacteroides thetaiotaomicron during murine gut colonization

doi: 10.1016/j.celrep.2023.113009

Figure Lengend Snippet: (A–E) Representative cross-sections of distal mouse colon fixed in Carnoy’s solution at (A) day 1 (D1), (B) D2, (C) D4, (D) D7, and (E) D42 of colonization by Bt . Blue, DAPI staining in the epithelium; red, DAPI staining in the lumen of gut, including bacteria, debris, and shed host nuclei; green, antibody staining for MUC2. Scale bars, 10 μm. (F) Mean Bt epithelial proximity. n = 3–4/group, t test, *p < 0.05.

Article Snippet: Rabbit polyclonal anti-mouse Muc2 , Santa Cruz Biotechnology , Cat#sc-15334.

Techniques: Staining, Bacteria

Journal: Cell reports

Article Title: Dynamic genetic adaptation of Bacteroides thetaiotaomicron during murine gut colonization

doi: 10.1016/j.celrep.2023.113009

Figure Lengend Snippet:

Article Snippet: Rabbit polyclonal anti-mouse Muc2 , Santa Cruz Biotechnology , Cat#sc-15334.

Techniques: Virus, Mutagenesis, Isolation, Recombinant, Saline, Enzyme-linked Immunosorbent Assay, SYBR Green Assay, cDNA Synthesis, Shotgun Sequencing, Sequencing, Software, Spectrophotometry, Mass Spectrometry, Microscopy

Journal: Cell reports

Article Title: Dynamic genetic adaptation of Bacteroides thetaiotaomicron during murine gut colonization

doi: 10.1016/j.celrep.2023.113009

Figure Lengend Snippet: (A–E) Representative cross-sections of distal mouse colon fixed in Carnoy’s solution at (A) day 1 (D1), (B) D2, (C) D4, (D) D7, and (E) D42 of colonization by Bt . Blue, DAPI staining in the epithelium; red, DAPI staining in the lumen of gut, including bacteria, debris, and shed host nuclei; green, antibody staining for MUC2. Scale bars, 10 μm. (F) Mean Bt epithelial proximity. n = 3–4/group, t test, *p < 0.05.

Article Snippet: After deparaffinization and rehydration, slides were incubated in lysozyme solution at 37°C for 20 min, and then in antigen retrieval solution (10 mM sodium citrate [pH 6.0]) at 90°C for 10 min. For mucus visualization, a polyclonal rabbit anti-mouse Muc2-specific antibody (Santa Cruz Biotechnology) was diluted 1:100 in blocking buffer (Dako), applied to the slide, and incubated for 2 h in the dark at room temperature.

Techniques: Staining, Bacteria

Journal: Cell reports

Article Title: Dynamic genetic adaptation of Bacteroides thetaiotaomicron during murine gut colonization

doi: 10.1016/j.celrep.2023.113009

Figure Lengend Snippet:

Article Snippet: After deparaffinization and rehydration, slides were incubated in lysozyme solution at 37°C for 20 min, and then in antigen retrieval solution (10 mM sodium citrate [pH 6.0]) at 90°C for 10 min. For mucus visualization, a polyclonal rabbit anti-mouse Muc2-specific antibody (Santa Cruz Biotechnology) was diluted 1:100 in blocking buffer (Dako), applied to the slide, and incubated for 2 h in the dark at room temperature.

Techniques: Virus, Mutagenesis, Isolation, Recombinant, Saline, Enzyme-linked Immunosorbent Assay, SYBR Green Assay, cDNA Synthesis, Shotgun Sequencing, Sequencing, Software, Spectrophotometry, Mass Spectrometry, Microscopy

Journal: Experimental & Molecular Medicine

Article Title: ERdj5 protects goblet cells from endoplasmic reticulum stress-mediated apoptosis under inflammatory conditions

doi: 10.1038/s12276-023-00945-x

Figure Lengend Snippet: a Periodic acid Schiff (PAS) staining of the colons of WT and ERdj5- KO mice (×100, scale bar 500 µm; n = 6 per group). b Goblet cell count per crypt in PAS images ( n = 15–25 per group). c Representative immunofluorescence images of MUC2 (green) in colon tissues, with DAPI (blue) for nuclear staining (×200, scale bar 200 µm). d Level of MUC2 mRNA expression ( n = 6–8 per group). e Representative immunofluorescence images of colonoids originating from WT or ERdj5- KO mice treated with vehicle or Pam 3 CSK 4 . MUC2 (green), TUNEL (red), E-cadherin (magenta), and DAPI (blue) (left panel, ×200 and right panel, ×800). f MUC2 + cell counts per high-power field (HPF) and the percentage of TUNEL-positive cells among MUC2 + and MUC2 - cells determined in colonoid images ( n = 5 per group). g Representative immunofluorescence images of colon tissues from WT or ERdj5- KO mice on Day 2 following DSS treatment. CLCA1 (red), TUNEL (green), and DAPI (blue) (left panel, ×200 and right panel, ×630). h CLCA1 + cell count per HPF and TUNEL-positive percentages among CLCA1 + cells from colon tissue images ( n = 4 per group). e , g Scale bars correspond to 100 μm and 20 μm, respectively. The data are representative of three independent experiments, and the values are expressed as the mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001; one-way ANOVA followed by Tukey’s test.

Article Snippet: The samples were stained with primary antibodies, including rabbit anti-mouse MUC2 (PA5-79702, 1:200; Invitrogen), rabbit anti-mouse CLCA1 (clone EPR12254-88, 1:200; Abcam), Alexa Fluor 647-conjugated rat anti-mouse E-cadherin (clone DECMA-1, 1:200; BioLegend), rabbit anti-mouse ZO-1 (61-7300, 1:250; Invitrogen), mouse anti-mouse Claudin-1 (clone 2H10D10, 1:250; Invitrogen), rabbit anti-mouse NF-κB (clone D14E12, 1:500; Cell Signaling Technology), rabbit anti-mouse myeloperoxidase (MPO; PA5-16672, 1:200; Invitrogen), APC-conjugated Ly6G (clone 1A8, 1:200; BD Biosciences), and goat anti-mouse lysozyme C (sc-27958, 1:200; Santa Cruz Biotechnology, Dallas, TX, USA).

Techniques: Staining, Cell Counting, Immunofluorescence, Expressing, TUNEL Assay

Journal: Experimental & Molecular Medicine

Article Title: ERdj5 protects goblet cells from endoplasmic reticulum stress-mediated apoptosis under inflammatory conditions

doi: 10.1038/s12276-023-00945-x

Figure Lengend Snippet: WT and ERdj5- KO mice were administered 2% DSS in drinking water for 5 days and subsequently provided normal water. The mice were orally administered 500 mg/kg UDCA daily ( n = 3–5 per group). a Bodyweight. b Colon length. c Representative H&E-stained colon tissue images (×100, scale bar 100 µm). Thin arrows indicate hyperplasia in affected areas. Thick arrows indicate multifocal inflammatory cell infiltration; m, mucosa; sm, submucosa; mm, muscular layer. d Histological scoring. e GRP78 mRNA expression in colon tissues. f CXCL1, g IL-1 and IL-6 levels in colon homogenates. h Zo-1 and Cldn1 mRNA levels in colon tissues. i Representative PAS staining (×100) and MUC2 immunofluorescence (×200) images of the colon. Scale bar corresponds to 200 µm. j MUC2 mRNA expression. The data are representative of three independent experiments, and the values are expressed as the mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001; ns, not significant; one-way ANOVA was followed by Tukey’s test.

Article Snippet: The samples were stained with primary antibodies, including rabbit anti-mouse MUC2 (PA5-79702, 1:200; Invitrogen), rabbit anti-mouse CLCA1 (clone EPR12254-88, 1:200; Abcam), Alexa Fluor 647-conjugated rat anti-mouse E-cadherin (clone DECMA-1, 1:200; BioLegend), rabbit anti-mouse ZO-1 (61-7300, 1:250; Invitrogen), mouse anti-mouse Claudin-1 (clone 2H10D10, 1:250; Invitrogen), rabbit anti-mouse NF-κB (clone D14E12, 1:500; Cell Signaling Technology), rabbit anti-mouse myeloperoxidase (MPO; PA5-16672, 1:200; Invitrogen), APC-conjugated Ly6G (clone 1A8, 1:200; BD Biosciences), and goat anti-mouse lysozyme C (sc-27958, 1:200; Santa Cruz Biotechnology, Dallas, TX, USA).

Techniques: Staining, Expressing, Immunofluorescence

Journal: Scientific reports

Article Title: Prevention of Shiga toxin 1-caused colon injury by plant-derived recombinant IgA.

doi: 10.1038/s41598-022-22851-4

Figure Lengend Snippet: Figure 4. Activation of caspase-3 and depletion of secretory granules in the colon epithelium upon intrarectal exposure to Stx1. Frozen sections (a,c) or paraffin sections (b) of colon tissue from Stx1-exposed or control (PBS) mice were stained with anti-activated caspase-3 antibodies (red in a), Alcian blue dye (blue in b), or anti- MUC2 antibodies (red in c). Nuclei were visualized by staining with DAPI (blue in a,c), and with hematoxylin (violet in b). Arrows indicate signals from activated caspase-3 (a). Fixation of tissues was performed with formalin-PBS (a,c) or Carnoy’s fixative (b). Bars represent 50 μm. Each mouse was exposed to 100 ng of Stx1. (d) Quantification of secretory granules in the colon tissues in response to the amount of Stx1 (abscissa) exposed to each mouse. The ordinate indicates the percentage of the MUC2-positive area divided by the total area of the mucous epithelium (mean ± SEM) from four independent mice. There were no exclusions of data points.

Article Snippet: Restriction enzymes NdeI, SacI-HF, NotI-HF, SacII and NsiI-HF were purchased from New England BioLabs (Ipswich, MA, USA); SmaI, KpnI, SalI, binary vector pRI201-AN, Klenow fragment, a DNA Ligation Kit and a DNA Ligation Kit from Takara Bio (Shiga, Japan); HindIII from Nippon Gene (Tokyo, Japan); a protease inhibitor cocktail for plant cell and tissue extracts, myeloma proteins TEPC 15 (mouse IgA, κ) and MOPC 21 (mouse IgG1, κ), and Alcian Blue 8GX from Sigma-Aldrich (St. Louis, MO, USA); 2,2’-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), Mayer’s Hematoxylin Solution, Murashige and Skoog (MS) plant salt mixture, and gellan gum from Wako Pure Chemical Industries, Ltd. 10 Vol:. (1234567890) Scientific Reports | (2022) 12:17999 | https://doi.org/10.1038/s41598-022-22851-4 (Osaka, Japan); Vero cells (an African Green Monkey kidney-derived cell line) from the American Type Culture Collection (Manassas, VA, USA); fetal bovine serum (FBS) from Hyclone (South Logan, UT, USA); a Cell Counting Kit-8 from DOJINDO (Kumamoto, Japan); 4,6-Diamidino-2-phenylindole Dihydrochloride (DAPI) from Nacalai Tesque (Kyoto, Japan); a PierceTM BCA Protein Assay Kit, HiMarkTM Pre-Stained Protein Standards, MagicMarkTM XP Western Protein Standards, SuperSignalTM West Pico PLUS Chemiluminescent Substrate, Medium 199 and Alexa594-goat anti-rabbit IgG from Thermo Fisher Scientific (Waltham, MA, USA); goat anti-mouse κ, horseradish peroxidase (HRP)-goat anti-mouse IgA, HRP-goat anti-mouse IgG, HRP-donkey anti-goat IgG and HRP-goat anti-rabbit IgG from Southern Biotech (Birmingham, AL, USA); rabbit anti-mouse J chain from Proteintech (Wuhan, China); rabbit anti-human/mouse activated-caspase 3 from BD Biosciences (Franklin Lakes, NJ, USA); and rabbit anti-mouse MUC2 from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Activation Assay, Control, Staining

Journal: Scientific reports

Article Title: Prevention of Shiga toxin 1-caused colon injury by plant-derived recombinant IgA.

doi: 10.1038/s41598-022-22851-4

Figure Lengend Snippet: Figure 5. Prevention of Stx1-caused colon injury by an IgA plantibody specific for Stx1. Sixty ng of Stx1 was treated with the leaf extract of an IgA transgenic plant (Stx1 + IgA-Tg) containing 8 μg IgA (1.6 mg total proteins), that of a wild-type plant (Stx1 + WT) containing 1.6 mg of total protein, or PBS (indicated as Stx1 only) for 1 h. Mice were then exposed to each sample containing Stx1 or to PBS alone (indicated as PBS) through intrarectal administration. After 16 h, colon tissues were collected and embedded in O.C.T. compound. Frozen sections of the colon tissues were stained with anti-activated caspase-3 (a) or anti-MUC2 (b,c) antibodies. Nuclei were counterstained with DAPI (a,b). Arrows indicate signals from activated caspase-3 (a). Bars represent 50 μm. The percentages of the MUC2-positive area divided by the total area of the mucous epithelium are plotted (c). Horizontal bars represent the means for individual groups and error bars are SD (n = 4). There were no exclusions of data points. Statistical significance was analyzed by ANOVA followed by the Tukey test. **P < 0.01, ****P < 0.0001, ns P > 0.05.

Article Snippet: Restriction enzymes NdeI, SacI-HF, NotI-HF, SacII and NsiI-HF were purchased from New England BioLabs (Ipswich, MA, USA); SmaI, KpnI, SalI, binary vector pRI201-AN, Klenow fragment, a DNA Ligation Kit and a DNA Ligation Kit from Takara Bio (Shiga, Japan); HindIII from Nippon Gene (Tokyo, Japan); a protease inhibitor cocktail for plant cell and tissue extracts, myeloma proteins TEPC 15 (mouse IgA, κ) and MOPC 21 (mouse IgG1, κ), and Alcian Blue 8GX from Sigma-Aldrich (St. Louis, MO, USA); 2,2’-Azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS), Mayer’s Hematoxylin Solution, Murashige and Skoog (MS) plant salt mixture, and gellan gum from Wako Pure Chemical Industries, Ltd. 10 Vol:. (1234567890) Scientific Reports | (2022) 12:17999 | https://doi.org/10.1038/s41598-022-22851-4 (Osaka, Japan); Vero cells (an African Green Monkey kidney-derived cell line) from the American Type Culture Collection (Manassas, VA, USA); fetal bovine serum (FBS) from Hyclone (South Logan, UT, USA); a Cell Counting Kit-8 from DOJINDO (Kumamoto, Japan); 4,6-Diamidino-2-phenylindole Dihydrochloride (DAPI) from Nacalai Tesque (Kyoto, Japan); a PierceTM BCA Protein Assay Kit, HiMarkTM Pre-Stained Protein Standards, MagicMarkTM XP Western Protein Standards, SuperSignalTM West Pico PLUS Chemiluminescent Substrate, Medium 199 and Alexa594-goat anti-rabbit IgG from Thermo Fisher Scientific (Waltham, MA, USA); goat anti-mouse κ, horseradish peroxidase (HRP)-goat anti-mouse IgA, HRP-goat anti-mouse IgG, HRP-donkey anti-goat IgG and HRP-goat anti-rabbit IgG from Southern Biotech (Birmingham, AL, USA); rabbit anti-mouse J chain from Proteintech (Wuhan, China); rabbit anti-human/mouse activated-caspase 3 from BD Biosciences (Franklin Lakes, NJ, USA); and rabbit anti-mouse MUC2 from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Techniques: Transgenic Assay, Staining